Method for simultaneously extracting rhaponticin, deoxyrhaponticin and anthraquinone compounds from rheum officinale

Abstract

The invention provides a method for simultaneously extracting rhaponticin, deoxyrhaponticin and anthraquinone compounds from rheum officinale, which can solve the problems of resource waste, high cost, complex process and low recovery rate caused by the fact that rhaponticin, deoxyrhaponticin and anthraquinone compounds cannot be simultaneously obtained by the existing method. The method comprises the following steps: cooking raw materials with boiling water to obtain a first filtrate and residues; adding industrial ammonia water and a first alcohol solution into the residues, stirring, extracting and filtering to obtain a second filtrate, concentrating, refrigerating and filtering to obtain a third filtrate and a first precipitate; modifying macroporous adsorption resin; dissolving the first precipitate with a small-polarity organic solvent, filtering to obtain a fourth filtrate and a second precipitate, concentrating, centrifuging, and combining a clear liquid with the first filtrate and the third filtrate to obtain a mixed liquid; adsorbing the mixed solution with modified macroporous adsorption resin, washing with water, eluting, concentrating and drying to obtain the anthraquinone compound; dissolving the second precipitate, filtering to obtain a fifth filtrate and a third precipitate, and concentrating and crystallizing to obtain a rhaponticin crude product; and refining.

Description

Technical Field

[0001] The invention relates to a method for simultaneously extracting rhubarb glycoside, deoxyrhubarb glycoside and anthraquinone compounds from plants, and in particular to a method for simultaneously extracting rhubarb glycoside, deoxyrhubarb glycoside and anthraquinone compounds from rhubarb. Background Art

[0002] Rhubarb is a variety included in the 2020 edition of the Pharmacopoeia of the People's Republic of China. It is the rhizome of Rheum palmatum, Rheum tanguticum, and Rheum officinale, Rheum palmatum, Rheum genus, Rheum palmatum, Polygonaceae. As early as the beginning of the 20th century, researchers studied the chemical composition of different types of rhubarb. The results showed that there are about 200 chemical components in rhubarb, mainly rhubarb anthraquinone compounds, rhubarb anthrone compounds, rhubarb tannin compounds, rhubarb stilbene compounds, and other chemical components such as rhubarb volatile oil compounds, rhubarb polysaccharide compounds, inorganic acids and organic acids.

[0003] According to the provisions of the pharmacopoeia, authentic rhubarb (palm leaf rhubarb, Tanggut rhubarb, medicinal rhubarb) all contain sennosides, and the roots and rhizomes of rhubarb plants such as North China rhubarb, Hetao rhubarb, and wave-leaved rhubarb are used as medicine in some areas or among the people. The characteristic components contained in these rhubarbs are diphenyl glycoside compounds, with rhubarb glycoside and deoxyrhubarb glycoside as representative components. In addition, they also contain combined anthraquinones, polysaccharides and other components, and have anti-tumor, anti-inflammatory, antioxidant, anti-pulmonary fibrosis, anti-thrombotic, and anti-Alzheimer's disease properties. In addition, the product of rhubarb glycoside after intestinal metabolism, danye rhubarb, also has good activity in anti-inflammatory and anti-thrombotic aspects. Therefore, it is of great application prospect to develop an industrial production method for simultaneously extracting rhubarb glycoside, deoxyrhubarb glycoside and anthraquinone compounds from rhubarb plants.

[0004] Chinese patent with publication number CN102887925B discloses a method for extracting rhubarb glycoside from Hetao rhubarb, which adopts ethanol extraction, petroleum ether / ethyl acetate mixed solvent extraction and degreasing, water phase filtration to obtain filter cake, ethanol recrystallization, and finally rhubarb glycoside is obtained, with a recovery rate of more than 70%.

[0005] The Chinese patent with publication number CN103421058B discloses a method for separating and purifying deoxyrhubarb glycosides with high efficiency and precision. The concentrated extract of Lhasa rhubarb is dissolved in water, purified twice with macroporous resin, and then recrystallized to obtain deoxyrhubarb glycosides.

[0006] A Chinese patent with publication number CN105330707B discloses an industrial production technology for simultaneously separating rhubarb glycoside and deoxyrhubarb glycoside. The steps are as follows: ethanol extraction of raw materials, concentration, polyamide resin separation, and ethanol gradient elution to obtain rhubarb glycoside and deoxyrhubarb glycoside respectively, and the mother liquor is repeatedly subjected to polyamide separation, with a final recovery rate of more than 90%.

[0007] The Chinese patent with publication number CN102702283B discloses a method for quickly separating and preparing high-purity deoxyrhubarb glycoside and rhubarb glycoside, and the method comprises the following steps: extracting the raw materials with alcohol and drying them to obtain an extract; mixing four solvents, methanol, water, n-butanol and chloroform, and shaking and standing them for stratification, and collecting the upper phase solvent and the lower phase solvent; dissolving the extract in the lower phase solvent, and separating by high-speed countercurrent chromatography, so as to prepare rhubarb glycoside and deoxyrhubarb glycoside, respectively.

[0008] A Chinese patent with publication number CN102786563A discloses a preparation method for separating three diphenylethylene glycoside monomer compounds from rhubarb. The method comprises the following steps: extracting by reflux with ethanol, concentrating, extracting with petroleum ether and ethyl acetate in sequence, applying the aqueous phase to a non-polar macroporous resin column, eluting with ethanol, and drying to obtain a crude diphenylethylene glycoside extract; and then separating by high-speed countercurrent chromatography to obtain three diphenylethylene glycoside substances, namely, trans-rhubarb glycoside, cis-rhubarb glycoside, and trans-deoxyrhubarb glycoside.

[0009] However, the problems existing in the above-mentioned existing preparation methods are: since it is impossible to obtain rhubarb glycoside, deoxyrhubarb glycoside and anthraquinone compounds at the same time, it leads to waste of resources and high production cost; the process is complicated and the cycle is long; and the recovery rate is low. Summary of the invention

[0010] The purpose of the present invention is to solve the technical problems that the existing preparation method cannot simultaneously obtain rhubarb glycoside, deoxyrhubarb glycoside and anthraquinone compounds, resulting in waste of resources, high production cost, complex process and low recovery rate, and provide a method for simultaneously extracting rhubarb glycoside, deoxyrhubarb glycoside and anthraquinone compounds from rhubarb.

[0011] In order to achieve the above object, the present invention adopts the following technical solution:

[0012] A method for simultaneously extracting rhubarb glycoside, deoxyrhubarb glycoside and anthraquinone compounds from rhubarb, which is special in that it comprises the following steps:

[0013] Step 1, crushing the rhubarb raw material, and then placing it in boiling water for steaming, filtering, and collecting the first filtrate and the residue respectively;

[0014] Step 2, adding industrial ammonia water and the first alcohol solution to the residue, stirring and extracting, filtering to obtain a second filtrate, and then sequentially concentrating the second filtrate, refrigerating and placing it, filtering, and collecting a third filtrate and a first precipitate respectively; the mass of the industrial ammonia water is 0.5-2% of the mass of the residue; the mass concentration of the first alcohol solution is 80-95%;

[0015] Step 3: Extracting anthraquinone compounds, crude rhubarb glycosides and crude deoxyrhubarb glycosides

[0016] Step 3.1, selecting a macroporous adsorption resin and modifying it with ethylene glycol and industrial ammonia water;

[0017] Step 3.2, placing the first precipitate collected in step 2 in a low-polarity organic solvent, stirring and dissolving, filtering, collecting the fourth filtrate and the second precipitate respectively, and then concentrating and centrifuging the fourth filtrate in turn, combining the clear liquid obtained by centrifugation with the first filtrate collected in step 1 and the third filtrate collected in step 2 to obtain a mixed liquid;

[0018] Step 3.3, loading the modified macroporous adsorption resin in step 3.1 into a chromatographic column, adsorbing the mixed solution, and then washing with water and eluting with a second alcohol solution in sequence, collecting the eluate, concentrating it and spray drying it to obtain anthraquinone compounds; placing the second precipitate collected in step 3.2 into a third alcohol solution, stirring and dissolving it, filtering it, collecting the fifth filtrate and the third precipitate respectively, and then concentrating the fifth filtrate, leaving it for crystallization, to obtain a crude product of rhubarb glycoside, wherein the third precipitate is a crude product of deoxyrhubarb glycoside;

[0019] Step 4: purify the crude rhubarb glycoside and the crude deoxyrhubarb glycoside respectively to obtain rhubarb glycoside and deoxyrhubarb glycoside respectively, and complete the extraction of rhubarb glycoside, deoxyrhubarb glycoside and anthraquinone compounds.

[0020] Furthermore, step 3.1 is specifically as follows:

[0021] A macroporous adsorption resin is selected, and the macroporous adsorption resin, ethylene glycol and industrial ammonia water are mixed in a mass volume ratio of 1:5-8:0.3-0.6, reacted at 60-80°C for 5-10 hours, then allowed to stand for 30-50 minutes, filtered, washed with water to neutrality and dried to obtain a modified macroporous adsorption resin.

[0022] Further, in step 2, industrial ammonia water and the first alcohol solution are added to the residue, and the mixture is stirred and extracted for 2-3 times, each time for 1-3 hours, and filtered to obtain a second filtrate, and then the second filtrate is first concentrated to a specific gravity of 1.1-1.2, and then refrigerated for more than 12 hours, filtered, and the third filtrate and the first precipitate are respectively collected;

[0023] The mass of the first alcohol solution is 5-10 times the mass of the residue, and the first alcohol solution is a methanol solution or an ethanol solution.

[0024] Furthermore, in step 3.1, the macroporous adsorption resin is LX-38 resin or ADS-17 resin; in step 3.2, the mass of the small polar organic solvent is 3-5 times the mass of the first precipitate, and the small polar organic solvent is water-saturated n-hexane, water-saturated petroleum ether or water-saturated dichloromethane; the number of stirring and dissolving is 2-3 times, each time for 1-1.5 hours;

[0025] In step 3.3, the mass ratio of the modified macroporous adsorption resin to the rhubarb raw material is 1-2:1; the mass concentration of the second alcohol solution is 50-80%, and the dosage is 2-3 chromatographic column volumes; the mass concentration of the third alcohol solution is 10-30%, and the mass is 5-8 times the mass of the second precipitate; the stirring and dissolving times is 1-2 times, each time for 1-2 hours; the fifth filtrate is concentrated to a specific gravity of 1.15-1.25; the second alcohol solution and the third alcohol solution are methanol solutions or ethanol solutions.

[0026] Furthermore, step 2 is specifically as follows:

[0027] Add industrial ammonia water with a mass of 1% of the residue mass and ethanol solution with a mass concentration of 90% which is 8 times the mass of the residue to the residue, stir and extract twice, each time for 2 hours, filter to obtain the second filtrate, and then concentrate the second filtrate to a specific gravity of 1.15, refrigerate for 12 hours, filter, and collect the third filtrate and the first precipitate respectively.

[0028] Furthermore, step 3 is specifically as follows:

[0029] Step 3.1, select LX-38 resin, mix LX-38 resin, ethylene glycol and industrial ammonia water in a mass volume ratio of 1:6:0.5, react at 70°C for 8 hours, then stand for 40 minutes, filter, wash with water to neutrality and dry to obtain modified LX-38 resin;

[0030] Step 3.2, the first precipitate collected in step 2 is placed in water-saturated petroleum ether with a mass 4 times that of the first precipitate, stirred and dissolved twice, each time for 1.5 hours, filtered, and the fourth filtrate and the second precipitate are collected respectively, and then the fourth filtrate is concentrated and centrifuged in turn, and the clear liquid obtained by centrifugation is combined with the first filtrate collected in step 1 and the third filtrate collected in step 2 to obtain a mixed solution;

[0031] Step 3.3, loading the modified LX-38 resin with a mass of 1 times that of the rhubarb raw material into a chromatographic column, adsorbing the mixed solution, then washing with water until the liquid is clear, eluting with an ethanol solution with a mass concentration of 70% and an amount of 3 chromatographic column volumes, collecting the eluate, concentrating and spray-drying to obtain anthraquinone compounds; placing the second precipitate collected in step 3.2 into an ethanol solution with a mass concentration of 20% that is 6 times that of the second precipitate, stirring and dissolving it twice, each time for 1 hour, filtering, collecting the fifth filtrate and the third precipitate respectively, and then concentrating the fifth filtrate to a specific gravity of 1.20, leaving it for crystallization, to obtain crude rhubarb glycoside, and the third precipitate is crude deoxyrhubarb glycoside.

[0032] Furthermore, step 1 is specifically as follows:

[0033] The rhubarb raw material is crushed into 40-60 meshes, then placed in boiling water with a mass 3-5 times that of the rhubarb raw material and boiled for 5-10 minutes, filtered, and the first filtrate and the residue are collected respectively; the rhubarb raw material is the root of wavy leaf rhubarb.

[0034] Furthermore, step 4 is specifically as follows:

[0035] At a temperature of 55-65° C., using a fourth alcohol solution with a mass concentration of 50-60% and a mass 10-15 times that of the crude rhubarb glycoside to dissolve the crude rhubarb glycoside, then using activated carbon with a mass of 2-5% of the mass of the crude rhubarb glycoside to decolorize, filtering, and allowing to crystallize to obtain rhubarb glycoside; the fourth alcohol solution is a methanol solution or an ethanol solution;

[0036] At a temperature of 55-65°C, ethyl acetate or acetone in an amount 8-15 times the mass of the crude deoxyrhubarb glycoside is used to dissolve the crude deoxyrhubarb glycoside, and then filtered and allowed to crystallize to obtain deoxyrhubarb glycoside, thereby completing the extraction of rhubarb glycoside, deoxyrhubarb glycoside and anthraquinone compounds.

[0037] Furthermore, step 1 is specifically as follows:

[0038] The root of Rheum officinale was crushed into 40-60 mesh, and then placed in boiling water with a mass 5 times that of the root of Rheum officinale and boiled for 8 minutes, filtered, and the first filtrate and the residue were collected separately.

[0039] Furthermore, step 4 is specifically as follows:

[0040] At a temperature of 60° C., using an ethanol solution with a mass concentration of 60% and a mass 12 times that of the crude rhubarb glycoside to dissolve the crude rhubarb glycoside, and then using activated carbon with a mass of 3% of the mass of the crude rhubarb glycoside to decolorize it, filtering it, and allowing it to crystallize to obtain rhubarb glycoside;

[0041] At a temperature of 60°C, ethyl acetate 10 times the mass of the crude deoxyrhubarb glycoside was used to dissolve the crude deoxyrhubarb glycoside, and then filtered and allowed to crystallize to obtain deoxyrhubarb glycoside, thereby completing the extraction of rhubarb glycoside, deoxyrhubarb glycoside and anthraquinone compounds.

[0042] Beneficial effects of the present invention:

[0043] 1. The method provided by the present invention for simultaneously extracting rhubarb glycoside, deoxyrhubarb glycoside and anthraquinone compounds from rhubarb has a simple process route and low production cost, can avoid the biological decomposition of rhubarb glycoside and deoxyrhubarb glycoside, improves the recovery rate, is conducive to the comprehensive development of raw materials, and is more suitable for industrial operation. Since rhubarb itself carries a biological enzyme that can enzymatically hydrolyze glycosidic bonds, the biological enzyme can directly act on rhubarb glycoside, deoxyrhubarb glycoside and rhubarb anthraquinone glycoside to enzymatically hydrolyze them into sugars and aglycones, resulting in reduced content and increased extraction costs. Therefore, in order to improve the extraction rate and avoid the degradation of rhubarb glycoside, deoxyrhubarb glycoside and anthraquinone compounds, the present invention pre-treats the raw materials before extraction and quickly inactivates the enzyme by boiling water, thereby avoiding the enzymatic hydrolysis of rhubarb glycoside, deoxyrhubarb glycoside and anthraquinone compounds during the extraction process, thereby improving the extraction rate; at the same time, the method can be used to extract and separate some chemical components such as anthraquinones and tannins, which is beneficial to the separation and purification of rhubarb glycoside and deoxyrhubarb glycoside in the later stage, and the filtration difficulty caused by tannin precipitation can be avoided in the alkali-alcohol extraction precipitation and filtration link.

[0044] 2. The present invention innovatively adopts alkaline alcohol solvent to extract rhubarb raw materials, uses high-concentration ethanol as the extraction solvent, and adds a weak alkaline solution of a specific concentration, which can greatly reduce the solubility of impurities. At the same time, it can also avoid the degradation of rhubarb glycoside and deoxyrhubarb glycoside caused by excessive alkalinity, laying a good foundation for the subsequent purification of rhubarb glycoside, deoxyrhubarb glycoside and anthraquinone compounds, and greatly shortening the subsequent purification process.

[0045] 3. Since the raw material of rhubarb is rich in anthraquinone compounds, has more phenolic hydroxyl groups, and has stronger polarity and hydrogen bond forming ability, in the present invention, amino functional groups are introduced based on the medium-polarity macroporous resin LX-38 or ADS-17 to enhance the original hydrophilicity and polarity of the resin, so that it forms stronger hydrogen bonds with anthraquinone compounds, has stronger adsorption for anthraquinone compounds, and is more conducive to the separation of such compounds.

[0046] 4. The present invention first uses a low-polarity organic solvent in combination with a modified macroporous adsorption resin and a second alcohol solution to extract anthraquinone compounds, and then, according to the physical and chemical properties of rhubarb glycoside and deoxyrhubarb glycoside, a low-concentration third alcohol solution with good selectivity for rhubarb glycoside and poor solubility for deoxyrhubarb glycoside is used for extraction, thereby achieving the separation of rhubarb glycoside and deoxyrhubarb glycoside, shortening the production cycle, and the process is simple, easy to operate, and more suitable for industrial production. BRIEF DESCRIPTION OF THE DRAWINGS

[0047] Figure 1 The HPLC spectrum of the root of Rheum officinale in Example 1 of the method for simultaneously extracting rheum officinale glycoside, deoxyrheum officinale glycoside and anthraquinone compounds from Rheum officinale of the present invention;

[0048] Figure 2 is the HPLC spectrum of the residue collected in Example 1 of the present invention;

[0049] Figure 3 This is the HPLC spectrum of rhubarb glycoside obtained in Example 1 of the present invention;

[0050] Figure 4 This is the HPLC spectrum of deoxyrhubarb glycoside obtained in Example 1 of the present invention. DETAILED DESCRIPTION

[0051] Embodiment 1

[0052] A method for simultaneously extracting rhubarb glycoside, deoxyrhubarb glycoside and anthraquinone compounds from rhubarb, characterized in that it comprises the following steps:

[0053] Step 1: Crush 100 kg of rhubarb roots to 40-60 mesh, then place in boiling water 5 times the mass of rhubarb roots and cook for 8 minutes, filter, and collect the first filtrate and the residue respectively. Figure 1 and Figure 2 The HPLC spectra of the root and residue of Rheum officinale are respectively Figure 1 and Figure 2 It can be seen that after the biological enzymes contained in the roots of Rhubarb were inactivated by steaming, the contents of rhubarb glycoside and deoxyrhubarb glycoside contained in it did not lose, but increased instead.

[0054] Step 2: add industrial ammonia water with a mass of 1% of the residue mass and ethanol solution with a mass concentration of 90% with a mass amount of 8 times the residue mass to the residue, stir and extract twice, each time for 2 hours, filter to obtain the second filtrate, and then concentrate the second filtrate to a specific gravity of 1.15, refrigerate for 12 hours, filter, and collect the third filtrate and the first precipitate respectively.

[0055] Step 3: Extracting anthraquinone compounds, crude rhubarb glycosides and crude deoxyrhubarb glycosides

[0056] Step 3.1, select LX-38 resin, mix LX-38 resin, ethylene glycol and industrial ammonia water in a mass volume ratio of 1:6:0.5, react at 70°C for 8 hours, then stand for 40 minutes, filter, wash with water to neutrality and dry to obtain modified LX-38 resin;

[0057] Step 3.2, the first precipitate collected in step 2 is placed in water-saturated petroleum ether of 4 times the mass of the first precipitate, stirred and dissolved twice, each time for 1.5 hours, filtered, the fourth filtrate and the second precipitate are collected respectively, the water-saturated petroleum ether is recovered, and then the fourth filtrate is concentrated and centrifuged in sequence, and the clear liquid obtained by centrifugation is combined with the first filtrate collected in step 1 and the third filtrate collected in step 2 to obtain a mixed solution;

[0058] Step 3.3, load the modified LX-38 resin with a mass of 1 times the amount of the root of the wave-leaved rhubarb into the chromatographic column, adsorb the mixed solution, then wash with water until the liquid is clear, elute with an ethanol solution with a mass concentration of 70% and a dosage of 3 chromatographic column volumes, collect the eluate, concentrate and spray dry, and obtain 4.161kg of anthraquinone compounds with a content of 55.27%; put the second precipitate collected in step 3.2 into an ethanol solution with a mass concentration of 20% and a mass concentration of 6 times the second precipitate, stir and dissolve twice, each time for 1h, filter, collect the fifth filtrate and the third precipitate respectively, and then concentrate the fifth filtrate to a specific gravity of 1.20, place and crystallize, and obtain the crude product of rhubarb glycoside, and the third precipitate is the crude product of deoxyrhubarb glycoside. In this step, the spray-dried substance is detected by UV ultraviolet spectrophotometer, and it can be known that the content of anthraquinone compounds in the substance is 55.27%.

[0059] Step 4, at a temperature of 60 ° C, using an ethanol solution with a mass concentration of 60% and a mass 12 times that of the crude rhubarb glycoside to dissolve the crude rhubarb glycoside, and then using activated carbon with a mass of 3% of the mass of the crude rhubarb glycoside to decolorize, filter, and allow to crystallize to obtain 4.290 kg of rhubarb glycoside with a content of 98.76%; in this step, the material after being allowed to crystallize is subjected to HPLC detection to obtain Figure 3 ,Depend on Figure 3 It can be seen that the substance is rhubarb glycoside, with a content of 98.76%;

[0060] At a temperature of 60°C, ethyl acetate 10 times the mass of the crude deoxyrhubarb was used to dissolve the crude deoxyrhubarb, and then filtered and allowed to crystallize to obtain 4.083 kg of deoxyrhubarb with a content of 98.39%, completing the extraction of rhubarb, deoxyrhubarb and anthraquinone compounds.

[0061] In this step, the material after crystallization was subjected to HPLC detection to obtain Figure 4 ,Depend on Figure 4 It can be seen that the substance is deoxyrhubarb glycoside, and the content is 98.39%.

[0062] Embodiment 2

[0063] Step 1: Crush 100 kg of rhubarb roots to 40-60 mesh, then place in boiling water 3 times the mass of rhubarb roots and cook for 10 minutes, filter, and collect the first filtrate and the residue respectively.

[0064] Step 2: add industrial ammonia water with a mass of 0.5% of the residue mass and ethanol solution with a mass concentration of 80% with a mass amount of 5 times the residue mass to the residue, stir and extract 3 times, each time for 1 hour, filter to obtain the second filtrate, and then concentrate the second filtrate to a specific gravity of 1.1, refrigerate for 12.5 hours, filter, and collect the third filtrate and the first precipitate respectively.

[0065] Step 3: Extracting anthraquinone compounds, crude rhubarb glycosides and crude deoxyrhubarb glycosides

[0066] Step 3.1, select LX-38 resin, mix LX-38 resin, ethylene glycol and industrial ammonia water in a mass volume ratio of 1:8:0.3, react at 80°C for 5 hours, then stand for 30 minutes, filter, wash with water to neutrality and dry to obtain modified LX-38 resin;

[0067] Step 3.2, the first precipitate collected in step 2 is placed in water-saturated n-hexane in an amount 3 times the mass of the first precipitate, stirred and dissolved for 3 times, each time for 1.5 hours, filtered, the fourth filtrate and the second precipitate are collected respectively, the water-saturated n-hexane is recovered, and then the fourth filtrate is concentrated and centrifuged in sequence, and the clear liquid obtained by centrifugation is combined with the first filtrate collected in step 1 and the third filtrate collected in step 2 to obtain a mixed solution;

[0068] Step 3.3, load the modified LX-38 resin with a mass of 1 times the amount of the root of the wavy leaf rhubarb into the chromatographic column, adsorb the mixed solution, then wash with water until the liquid is clear, elute with an ethanol solution with a mass concentration of 50% and an amount of 2 chromatographic column volumes, collect the eluate, concentrate and spray dry, and obtain 4.472kg of anthraquinone compounds with a content of 50.86%; put the second precipitate collected in step 3.2 into an ethanol solution with a mass concentration of 10% and a mass concentration of 8 times the second precipitate, stir and dissolve twice, each time for 1h, filter, collect the fifth filtrate and the third precipitate respectively, and then concentrate the fifth filtrate to a specific gravity of 1.15, place it for crystallization, and obtain a crude product of rhubarb glycoside, and the third precipitate is a crude product of deoxyrhubarb glycoside. In this step, the method for detecting the content of anthraquinone compounds in the spray-dried material is the same as the method in Example 1.

[0069] Step 4, at a temperature of 55° C., using an ethanol solution with a mass concentration of 60% and a mass 15 times that of the crude rhubarb glycoside to dissolve the crude rhubarb glycoside, and then using activated carbon with a mass of 5% of the mass of the crude rhubarb glycoside to decolorize, filter, and allow to crystallize to obtain 4.256 kg of rhubarb glycoside with a content of 98.23%;

[0070] At a temperature of 55°C, ethyl acetate 15 times the mass of the crude deoxyrhubarb was used to dissolve the crude deoxyrhubarb, and then filtered and allowed to crystallize to obtain 4.027 kg of deoxyrhubarb with a content of 98.50%, completing the extraction of rhubarb, deoxyrhubarb and anthraquinone compounds.

[0071] Embodiment 3

[0072] Step 1: Crush 100 kg of rhubarb roots to 40-60 mesh, then place in boiling water 5 times the mass of rhubarb roots and cook for 5 minutes, filter, and collect the first filtrate and the residue respectively.

[0073] Step 2: Add 2% of the mass of the residue to the residue industrial ammonia water and 95% ethanol solution with a mass concentration of 10 times the mass of the residue, stir and extract twice, each time for 3 hours, filter to obtain the second filtrate, and then concentrate the second filtrate to a specific gravity of 1.2, refrigerate for 13 hours, filter, and collect the third filtrate and the first precipitate respectively.

[0074] Step 3: Extracting anthraquinone compounds, crude rhubarb glycosides and crude deoxyrhubarb glycosides

[0075] Step 3.1, select ADS-17 resin, mix ADS-17 resin, ethylene glycol and industrial ammonia water in a mass volume ratio of 1:5:0.6, react at 60°C for 10 hours, then let stand for 50 minutes, filter, wash with water to neutrality and dry to obtain modified ADS-17 resin;

[0076] Step 3.2, the first precipitate collected in step 2 is placed in water-saturated petroleum ether with a mass 5 times that of the first precipitate, stirred and dissolved twice, each time for 1 hour, filtered, and the fourth filtrate and the second precipitate are collected respectively, the water-saturated petroleum ether is recovered, and then the fourth filtrate is concentrated and centrifuged in sequence, and the clear liquid obtained by centrifugation is combined with the first filtrate collected in step 1 and the third filtrate collected in step 2 to obtain a mixed solution;

[0077] Step 3.3, the modified ADS-17 resin with a mass of 2 times the amount of the root of the wave-leaved rhubarb is loaded into the chromatographic column, the mixed solution is adsorbed, and then washed with water until the liquid is clear, eluted with an ethanol solution with a mass concentration of 80% and an amount of 3 chromatographic column volumes, the eluate is collected, concentrated and spray-dried to obtain 4.319kg of anthraquinone compounds with a content of 52.44%; the second precipitate collected in step 3.2 is placed in an ethanol solution with a mass concentration of 30% and stirred and dissolved once, each time for 2h, filtered, and the fifth filtrate and the third precipitate are collected respectively, and then the fifth filtrate is concentrated to a specific gravity of 1.25, and after crystallization, a crude product of rhubarb glycoside is obtained, and the third precipitate is a crude product of deoxyrhubarb glycoside. In this step, the method for detecting the content of anthraquinone compounds in the spray-dried material is the same as the method in Example 1.

[0078] Step 4, at a temperature of 65°C, using an ethanol solution with a mass concentration of 50% and a mass that is 10 times that of the crude rhubarb glycoside to dissolve the crude rhubarb glycoside, and then using activated carbon with a mass that is 2% of the mass of the crude rhubarb glycoside to decolorize, filter, and allow to crystallize to obtain 4.231 kg of rhubarb glycoside with a content of 98.19%;

[0079] At a temperature of 65°C, ethyl acetate in an amount 8 times the mass of the crude deoxyrhubarb was used to dissolve the crude deoxyrhubarb, and then filtered and allowed to crystallize to obtain 4.052 kg of deoxyrhubarb with a content of 98.55%, completing the extraction of rhubarb, deoxyrhubarb and anthraquinone compounds.

[0080] Embodiment 4

[0081] Step 1: Crush 100 kg of rhubarb roots to 40-60 mesh, then place in boiling water with a mass four times that of the rhubarb roots and cook for 9 minutes, filter, and collect the first filtrate and the residue respectively.

[0082] Step 2: add industrial ammonia water with a mass of 1.5% of the mass of the residue and ethanol solution with a mass concentration of 85% with a mass amount of 7 times the mass of the residue to the residue, stir and extract twice, each time for 1 hour, filter to obtain the second filtrate, and then concentrate the second filtrate to a specific gravity of 1.15, refrigerate for 12.5 hours, filter, and collect the third filtrate and the first precipitate respectively.

[0083] Step 3: Extracting anthraquinone compounds, crude rhubarb glycosides and crude deoxyrhubarb glycosides

[0084] Step 3.1, select LX-38 resin, mix LX-38 resin, ethylene glycol and industrial ammonia water in a mass volume ratio of 1:7:0.4, react at 75°C for 6 hours, then stand for 35 minutes, filter, wash with water to neutrality and dry to obtain modified LX-38 resin;

[0085] Step 3.2, the first precipitate collected in step 2 is placed in water-saturated dichloromethane with a mass 3 times that of the first precipitate, stirred and dissolved twice, each time for 1.2 hours, filtered, the fourth filtrate and the second precipitate are collected respectively, the water-saturated dichloromethane is recovered, and then the fourth filtrate is concentrated and centrifuged in sequence, and the clear liquid obtained by centrifugation is combined with the first filtrate collected in step 1 and the third filtrate collected in step 2 to obtain a mixed solution;

[0086] Step 3.3, load the modified LX-38 resin with a mass of 1.5 times the amount of the root of the wavy leaf rhubarb into the chromatographic column, adsorb the mixed solution, then wash with water until the liquid is clear, elute with an ethanol solution with a mass concentration of 75% and an amount of 2.5 chromatographic column volumes, collect the eluate, concentrate and spray dry, and obtain 4.380kg of anthraquinone compounds with a content of 51.83%; put the second precipitate collected in step 3.2 into an ethanol solution with a mass concentration of 15% and a mass concentration of 7 times the amount of the second precipitate, stir and dissolve once, each time for 1.5h, filter, collect the fifth filtrate and the third precipitate respectively, and then concentrate the fifth filtrate to a specific gravity of 1.20, place it for crystallization, and obtain a crude product of rhubarb glycoside, and the third precipitate is a crude product of deoxyrhubarb glycoside. In this step, the method for detecting the content of anthraquinone compounds in the spray-dried material is the same as the method in Example 1.

[0087] Step 4, at a temperature of 60°C, using an ethanol solution with a mass concentration of 55% and a mass 14 times that of the crude rhubarb glycoside to dissolve the crude rhubarb glycoside, and then using activated carbon with a mass of 4% of the mass of the crude rhubarb glycoside to decolorize, filter, and allow to crystallize to obtain 4.238 kg of rhubarb glycoside with a content of 98.46%;

[0088] At a temperature of 60°C, ethyl acetate 12 times the mass of the crude deoxyrhubarb was used to dissolve the crude deoxyrhubarb, and then filtered and allowed to crystallize to obtain 4.036 kg of deoxyrhubarb with a content of 98.09%, completing the extraction of rhubarb, deoxyrhubarb and anthraquinone compounds.

[0089] Embodiment 5

[0090] Step 1: Crush 100 kg of rhubarb roots to 40-60 mesh, then place in boiling water 5 times the mass of rhubarb roots and cook for 6 minutes, filter, and collect the first filtrate and the residue respectively.

[0091] Step 2: add industrial ammonia water with a mass of 0.8% of the residue mass and methanol solution with a mass concentration of 85% which is 6 times the mass of the residue to the residue, stir and extract twice, each time for 1 hour, filter to obtain the second filtrate, and then concentrate the second filtrate to a specific gravity of 1.15, refrigerate for 12 hours, filter, and collect the third filtrate and the first precipitate respectively.

[0092] Step 3: Extracting anthraquinone compounds, crude rhubarb glycosides and crude deoxyrhubarb glycosides

[0093] Step 3.1, select ADS-17 resin, mix ADS-17 resin, ethylene glycol and industrial ammonia water in a mass volume ratio of 1:8:0.3, react at 70°C for 8 hours, then let stand for 40 minutes, filter, wash with water to neutrality and dry to obtain modified ADS-17 resin;

[0094] Step 3.2, the first precipitate collected in step 2 is placed in water-saturated petroleum ether of 4 times the mass of the first precipitate, stirred and dissolved for 3 times, each time for 1.5 hours, filtered, the fourth filtrate and the second precipitate are collected respectively, the water-saturated petroleum ether is recovered, and then the fourth filtrate is concentrated and centrifuged in turn, and the clear liquid obtained by centrifugation is combined with the first filtrate collected in step 1 and the third filtrate collected in step 2 to obtain a mixed solution;

[0095] Step 3.3, the modified ADS-17 resin with a mass of 1 times the amount of the root of the wave-leaved rhubarb is loaded into the chromatographic column, the mixed solution is adsorbed, and then washed with water until the liquid is clear, and eluted with a methanol solution with a mass concentration of 60% and a dosage of 3 chromatographic column volumes, the eluate is collected, concentrated and spray-dried to obtain 4.257kg of anthraquinone compounds with a content of 53.07%; the second precipitate collected in step 3.2 is placed in a methanol solution with a mass concentration of 25% and stirred and dissolved twice, each time for 1h, filtered, and the fifth filtrate and the third precipitate are collected respectively, and then the fifth filtrate is concentrated to a specific gravity of 1.20, and after crystallization, a crude product of rhubarb glycoside is obtained, and the third precipitate is a crude product of deoxyrhubarb glycoside. In this step, the method for detecting the content of anthraquinone compounds in the spray-dried substance is the same as the method in Example 1.

[0096] Step 4, at a temperature of 60°C, using a methanol solution with a mass concentration of 55% and a mass 11 times that of the crude rhubarb glycoside to dissolve the crude rhubarb glycoside, and then using activated carbon with a mass of 3% of the mass of the crude rhubarb glycoside to decolorize, filter, and allow to crystallize to obtain 4.219 kg of rhubarb glycoside with a content of 98.42%;

[0097] At a temperature of 60°C, acetone 14 times the mass of the crude deoxyrhubarb was used to dissolve the crude deoxyrhubarb, and then filtered and allowed to crystallize to obtain 4.045 kg of deoxyrhubarb with a content of 98.25%, completing the extraction of rhubarb, deoxyrhubarb and anthraquinone compounds.

[0098] Comparative Example 1

[0099] The difference between this comparative example and Example 1 is that:

[0100] Step 1: Crush 100kg of rhubarb roots into 40-60 mesh.

[0101] Step 2: Add 8 times the amount of ethanol solution with a mass concentration of 90% and 1% ammonia water with a mass of the wavy leaf rhubarb roots to the crushed wavy leaf rhubarb roots, stir and extract twice, each time for 2 hours, filter, concentrate the filtrate to a specific gravity of 1.15, refrigerate for 12 hours, filter, and collect the filtrate and precipitate separately.

[0102] In step 3.3, 4.098 kg of anthraquinone compounds were obtained, with a content of 54.62%. In this step, the detection method was the same as in Example 1.

[0103] In step 4, 4.022 kg of rhubarb glycosides were obtained, with a content of 95.16%; and 3.835 kg of deoxyrhubarb glycosides were obtained, with a content of 93.28%.

[0104] Comparative Example 2

[0105] The difference between this comparative example and Example 1 is that:

[0106] Step 2: Add ethanol solution with a mass of 8 times that of the residue and a mass concentration of 90% to the residue, stir and extract twice, each time for 2 hours, filter to obtain the second filtrate, and then concentrate the second filtrate to a specific gravity of 1.15, refrigerate for 12 hours, filter, and collect the third filtrate and the first precipitate respectively.

[0107] In step 3.3, 4.307 kg of anthraquinone compounds were obtained, with a content of 48.12%. In this step, the detection method was the same as in Example 1.

[0108] In step 4, 4.506 kg of rhubarb glycosides were obtained, with a content of 88.73%; and 4.201 kg of deoxyrhubarb glycosides were obtained, with a content of 90.82%.

[0109] Comparative Example 3

[0110] The difference between this comparative example and Example 1 is that:

[0111] Without modifying the LX-38 resin, in step 3.3, LX-38 resin with a mass equal to 1 times that of the root of Rheum officinale was directly used to separate anthraquinone compounds, and 3.152 kg of anthraquinone compounds with a content of 40.73% were obtained. In this step, the detection method was the same as that in Example 1.

[0112] In step 4, 4.279 kg of rhubarb glycosides were obtained, with a content of 98.84%; and 4.095 kg of deoxyrhubarb glycosides were obtained, with a content of 98.18%.

[0113] In Examples 2 to 5 and Comparative Examples 2 to 3, the detection methods for the root and residue of Rheum officinale in step 1 and the two substances after crystallization in step 4 are the same as those in Example 1, and the corresponding HPLC spectra are similar to those in Example 1, which can be referred to Figures 1 to 4 .

[0114] In Comparative Example 1, the detection methods for the root of Rheum officinale in step 1 and the two substances after crystallization in step 4 are the same as those in Example 1, and the corresponding HPLC spectrum is similar to that in Example 1, which can be referred to Figure 1 , Figure 3 and Figure 4 .

[0115] The above is only a specific embodiment of the present invention, but the protection scope of the present invention is not limited thereto. Any changes or substitutions within the technical scope disclosed by the present invention should be included in the protection scope of the present invention. Therefore, the protection scope of the present invention should be based on the protection scope of the claims.

Claims

1. A method for simultaneously extracting rhubarb glycoside, deoxyrhubarb glycoside and anthraquinone compounds from rhubarb, characterized in that: The following steps are involved: Step 1, crushing the rhubarb raw material, and then placing it in boiling water for steaming, filtering, and collecting the first filtrate and the residue respectively; Step 2, adding industrial ammonia water and the first alcohol solution to the residue, stirring and extracting, filtering to obtain a second filtrate, and then sequentially concentrating the second filtrate, refrigerating and placing it, filtering, and collecting a third filtrate and a first precipitate respectively; the mass of the industrial ammonia water is 0.5-2% of the mass of the residue; the mass concentration of the first alcohol solution is 80-95%; Step 3: Extracting anthraquinone compounds, crude rhubarb glycosides and crude deoxyrhubarb glycosides Step 3.1, selecting a macroporous adsorption resin and modifying it with ethylene glycol and industrial ammonia water; Step 3.2, placing the first precipitate collected in step 2 in a low-polarity organic solvent, stirring and dissolving, filtering, collecting the fourth filtrate and the second precipitate respectively, and then concentrating and centrifuging the fourth filtrate in turn, combining the clear liquid obtained by centrifugation with the first filtrate collected in step 1 and the third filtrate collected in step 2 to obtain a mixed liquid; Step 3.3, loading the modified macroporous adsorption resin in step 3.1 into a chromatographic column, adsorbing the mixed solution, and then washing with water and eluting with a second alcohol solution in sequence, collecting the eluate, concentrating it and spray drying it to obtain anthraquinone compounds; The second precipitate collected in step 3.2 is placed in the third alcohol solution, stirred and dissolved, filtered, and the fifth filtrate and the third precipitate are collected respectively, and then the fifth filtrate is concentrated, and after standing for crystallization, a crude product of rhubarb glycoside is obtained, wherein the third precipitate is a crude product of deoxyrhubarb glycoside; Step 4: purify the crude rhubarb glycoside and the crude deoxyrhubarb glycoside respectively to obtain rhubarb glycoside and deoxyrhubarb glycoside respectively, and complete the extraction of rhubarb glycoside, deoxyrhubarb glycoside and anthraquinone compounds.

2. The method for simultaneously extracting rhubarb glycoside, deoxyrhubarb glycoside and anthraquinone compounds from rhubarb according to claim 1, characterized in that: Step 3.1 is as follows: A macroporous adsorption resin is selected, and the macroporous adsorption resin, ethylene glycol and industrial ammonia water are mixed in a mass volume ratio of 1:5-8:0.3-0.6, reacted at 60-80°C for 5-10 hours, then allowed to stand for 30-50 minutes, filtered, washed with water to neutrality and dried to obtain a modified macroporous adsorption resin.

3. The method for simultaneously extracting rhubarb glycoside, deoxyrhubarb glycoside and anthraquinone compounds from rhubarb according to claim 2, characterized in that: In step 2, industrial ammonia water and the first alcohol solution are added to the residue, and the mixture is stirred and extracted for 2-3 times, each time for 1-3 hours, and filtered to obtain a second filtrate, and then the second filtrate is first concentrated to a specific gravity of 1.1-1.2, and then refrigerated for more than 12 hours, filtered, and the third filtrate and the first precipitate are respectively collected; The mass of the first alcohol solution is 5-10 times the mass of the residue, and the first alcohol solution is a methanol solution or an ethanol solution.

4. The method for simultaneously extracting rhubarb glycoside, deoxyrhubarb glycoside and anthraquinone compounds from rhubarb according to claim 3, characterized in that: In step 3.1, the macroporous adsorption resin is LX-38 resin or ADS-17 resin; In step 3.2, the mass of the low-polarity organic solvent is 3-5 times the mass of the first precipitate, and the low-polarity organic solvent is water-saturated n-hexane, water-saturated petroleum ether or water-saturated dichloromethane; the stirring and dissolving is performed 2-3 times, each time for 1-1.5 hours; In step 3.3, the mass ratio of the modified macroporous adsorption resin to the rhubarb raw material is 1-2:1; the mass concentration of the second alcohol solution is 50-80%, and the dosage is 2-3 chromatographic column volumes; the mass concentration of the third alcohol solution is 10-30%, and the mass is 5-8 times the mass of the second precipitate; the stirring and dissolving times is 1-2 times, each time for 1-2 hours; the fifth filtrate is concentrated to a specific gravity of 1.15-1.25; the second alcohol solution and the third alcohol solution are methanol solutions or ethanol solutions.

5. The method for simultaneously extracting rhubarb glycoside, deoxyrhubarb glycoside and anthraquinone compounds from rhubarb according to claim 4, characterized in that: Step 2 is as follows: Add industrial ammonia water with a mass of 1% of the residue mass and ethanol solution with a mass concentration of 90% which is 8 times the mass of the residue to the residue, stir and extract twice, each time for 2 hours, filter to obtain the second filtrate, and then concentrate the second filtrate to a specific gravity of 1.15, refrigerate for 12 hours, filter, and collect the third filtrate and the first precipitate respectively.

6. The method for simultaneously extracting rhubarb glycoside, deoxyrhubarb glycoside and anthraquinone compounds from rhubarb according to claim 5, characterized in that: Step 3 is as follows: Step 3.1, select LX-38 resin, mix LX-38 resin, ethylene glycol and industrial ammonia water in a mass volume ratio of 1:6:0.5, react at 70°C for 8 hours, then stand for 40 minutes, filter, wash with water to neutrality and dry to obtain modified LX-38 resin; Step 3.2, the first precipitate collected in step 2 is placed in water-saturated petroleum ether with a mass 4 times that of the first precipitate, stirred and dissolved twice, each time for 1.5 hours, filtered, and the fourth filtrate and the second precipitate are collected respectively, and then the fourth filtrate is concentrated and centrifuged in turn, and the clear liquid obtained by centrifugation is combined with the first filtrate collected in step 1 and the third filtrate collected in step 2 to obtain a mixed solution; Step 3.3, loading the modified LX-38 resin with a mass of 1 times that of the rhubarb raw material into a chromatographic column, adsorbing the mixed solution, then washing with water until the liquid is clear, eluting with an ethanol solution with a mass concentration of 70% and an amount of 3 chromatographic column volumes, collecting the eluate, concentrating and spray-drying to obtain anthraquinone compounds; placing the second precipitate collected in step 3.2 into an ethanol solution with a mass concentration of 20% that is 6 times that of the second precipitate, stirring and dissolving it twice, each time for 1 hour, filtering, collecting the fifth filtrate and the third precipitate respectively, and then concentrating the fifth filtrate to a specific gravity of 1.20, leaving it for crystallization, to obtain crude rhubarb glycoside, and the third precipitate is crude deoxyrhubarb glycoside.

7. The method for simultaneously extracting rhubarb glycoside, deoxyrhubarb glycoside and anthraquinone compounds from rhubarb according to claim 1, characterized in that: Step 1 is as follows: The rhubarb raw material is crushed into 40-60 meshes, then placed in boiling water with a mass 3-5 times that of the rhubarb raw material and boiled for 5-10 minutes, filtered, and the first filtrate and the residue are collected respectively; the rhubarb raw material is the root of wavy leaf rhubarb.

8. The method for simultaneously extracting rhubarb glycoside, deoxyrhubarb glycoside and anthraquinone compounds from rhubarb according to claim 7, characterized in that: Step 4 is as follows: At a temperature of 55-65° C., using a fourth alcohol solution with a mass concentration of 50-60% and a mass 10-15 times that of the crude rhubarb glycoside to dissolve the crude rhubarb glycoside, then using activated carbon with a mass of 2-5% of the mass of the crude rhubarb glycoside to decolorize, filtering, and allowing to crystallize to obtain rhubarb glycoside; the fourth alcohol solution is a methanol solution or an ethanol solution; At a temperature of 55-65°C, ethyl acetate or acetone in an amount 8-15 times the mass of the crude deoxyrhubarb glycoside is used to dissolve the crude deoxyrhubarb glycoside, and then filtered and allowed to crystallize to obtain deoxyrhubarb glycoside, thereby completing the extraction of rhubarb glycoside, deoxyrhubarb glycoside and anthraquinone compounds.

9. The method for simultaneously extracting rhubarb glycoside, deoxyrhubarb glycoside and anthraquinone compounds from rhubarb according to claim 8, characterized in that: Step 1 is as follows: The root of Rheum officinale was crushed into 40-60 mesh, and then placed in boiling water with a mass 5 times that of the root of Rheum officinale for 8 minutes, filtered, and the first filtrate and the residue were collected separately.

10. The method for simultaneously extracting rhubarb glycoside, deoxyrhubarb glycoside and anthraquinone compounds from rhubarb according to claim 9, characterized in that: Step 4 is as follows: At a temperature of 60° C., using an ethanol solution with a mass concentration of 60% and a mass 12 times that of the crude rhubarb glycoside to dissolve the crude rhubarb glycoside, and then using activated carbon with a mass of 3% of the mass of the crude rhubarb glycoside to decolorize it, filtering it, and allowing it to crystallize to obtain rhubarb glycoside; At a temperature of 60°C, ethyl acetate 10 times the mass of the crude deoxyrhubarb glycoside was used to dissolve the crude deoxyrhubarb glycoside, and then filtered and allowed to crystallize to obtain deoxyrhubarb glycoside, thereby completing the extraction of rhubarb glycoside, deoxyrhubarb glycoside and anthraquinone compounds.

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