Process optimization method for purifying emodin by using macroporous resin

Abstract

The invention provides a process optimization method for purifying emodin through macroporous resin, and belongs to the technical field of chemical purification processes. The method comprises the following steps: respectively mixing different pretreated macroporous resins with an emodin adsorbate solution, carrying out adsorption under a static test condition until saturation, carrying out chromatographic analysis on the solutions before and after adsorption, and calculating an interaction mechanism between emodin and the macroporous resins by adopting a molecular docking technology, molecular dynamics and binding energy. Screening to obtain macroporous resin with the best adsorption effect and purification conditions; the method comprises the following steps: adsorbing to be saturated by adopting macroporous resin with an optimal adsorption effect and an emodin adsorbate solution under a dynamic adsorption condition, firstly washing impurities with water, then desorbing with ethanol to obtain an eluent, and concentrating and crystallizing the eluent to obtain the high-purity emodin. The purity of the purified emodin reaches up to 99.8%, and an efficient, simple, convenient and stable method is provided for obtaining of high-purity emodin and industrial large-scale production of emodin.

Description

Technical Field

[0001] The invention relates to the technical field of chemical purification processes, and in particular to a process optimization method for purifying emodin with a macroporous resin. Background Art

[0002] Rhamnosin, a hydroxyanthraquinone compound, is chemically named 1,3,8-trihydroxy-6-methylanthraquinone. It has been used as a traditional medicine for over 2,000 years and is commonly found in over 800 traditional Chinese medicine preparations. It is abundant in the Polygonum cuspidatum, Leguminosae, and Rhamnaceae plant families. Rhamnosin exhibits diverse pharmacological effects, including anticancer, anti-inflammatory, antiviral, and antioxidant properties. Molecular docking has previously revealed the therapeutic targets and pharmacological mechanisms of rhamnosin in the treatment of COVID-19. Through continued research, rhamnosin has become a valuable ingredient in traditional Chinese medicine preparations and dietary supplements. However, rhamnosin is virtually insoluble in water but soluble in ethanol and alkaline solutions. Current purification methods for rhamnosin include organic solvent extraction, column chromatography, polyamide, dextran gel, and ion exchange resins, but the yield of pure rhamnosin is low. Among them, patent CN103073409A discloses a method of collecting the eluate and recovering the solvent after separation and purification with a macroporous resin to obtain a crude product of rhein, and then purifying the crude product by silica gel column chromatography to obtain a pure product of rhein. This patent uses organic solvents chloroform and methanol (chloroform: methanol = 7:3) for elution. Similarly, the methods mentioned in patents CN104447270A, CN109517855A, CN116354810A, etc. all contain toxic solvents and have high requirements for operators and equipment.

[0003] Currently, traditional emodin purification technologies struggle to meet market demand for high-purity products. For example, silica gel column chromatography suffers from poor adsorption selectivity and small single-shot processing capacity, resulting in low purification efficiency. While multiple crystallization methods can improve purity, they consume large amounts of solvent and are prone to product loss during the crystallization process, significantly increasing production costs. Furthermore, existing processes are sensitive to parameters such as temperature and pH, resulting in poor production stability and difficulty ensuring consistent product quality. Furthermore, the organic solvents used in some processes are highly toxic and carry a high risk of residual residues, posing health risks to operators and limiting the application of emodin in high-end applications such as pharmaceuticals. Macroporous adsorption resins (MARs) are functional polymer materials that, compared to other adsorbents, offer advantages such as structural diversity, low cost, excellent acid and alkali resistance, high porosity, high specific surface area, environmental friendliness, and long service life. However, their application in the purification of emodin is currently lacking, hindering the development of research on its separation and purification. Summary of the Invention

[0004] In view of the above shortcomings of the existing technology, the present invention provides a process optimization method for purifying emodin with a macroporous resin. The present invention systematically screens macroporous resins that are beneficial to the purification of emodin and optimizes the process conditions for purifying emodin with a macroporous resin, thereby providing an efficient, simple and stable method for obtaining high-purity emodin and for the industrial-scale production of emodin.

[0005] To achieve the above purpose, the specific technical solutions of the present invention are as follows:

[0006] The present invention provides a process optimization method for purifying emodin with a macroporous resin, comprising the following steps:

[0007] Different pretreated macroporous resins were mixed with emodin adsorbate solutions and adsorbed under static test conditions until saturation. The solutions before and after adsorption were then chromatographically analyzed. Molecular docking, molecular dynamics, and binding energy were used to calculate the interaction mechanism between emodin and the macroporous resins, and the optimal macroporous resin and purification conditions were screened.

[0008] The macroporous resin with the best adsorption effect and the emodin adsorbate solution are used for adsorption to saturation under dynamic adsorption conditions, and the impurities are first washed with water and then desorbed with ethanol to obtain an eluate. The eluate is concentrated and crystallized to obtain high-purity emodin.

[0009] The present invention systematically conducts chromatographic analysis of samples before and after adsorption using SD300, D101, X-5, H103, and SV300 macroporous adsorption resins to identify optimal emodin adsorption solution conditions. The optimal macroporous resin is screened by accurately measuring the peak values ​​and separation of the main components and impurities, and calculating the emodin adsorption capacity. Molecular docking is then used to predict the interaction between the main structure of the macroporous resin and the emodin molecules. Gaussian software is used to calculate the size of the emodin molecule using quantum chemistry theory and the specific surface area test (BET) to calculate the resin pore size. This theoretically predicts whether the macroporous resin meets adsorption requirements. The invention obtains an adsorption isotherm of emodin molecules on a resin surface through investigation of static adsorption thermodynamics and kinetics, determines the adsorption mode of emodin molecules on the surface of a macroporous adsorption resin by combining models such as Langmuir and Freundlich and thermodynamic parameters, fits the adsorption process through a pseudo-first-order kinetic equation, a pseudo-second-order kinetic equation and an intra-particle diffusion equation, analyzes the adsorption mechanism of the H103 macroporous resin and the diffusion behavior of emodin molecules within the particles, and obtains emodin with higher purity through dynamic adsorption and analytical technology, concentration, recrystallization and drying, thereby providing a scientific basis for the rational utilization of resources.

[0010] Preferably, the method for pretreating the macroporous resin is as follows: soaking the macroporous resin in ethanol to remove upper suspended particles, and then sequentially washing with water, acid washing, water washing, alkali washing, and water washing to obtain the pretreated macroporous resin.

[0011] Preferably, the different pretreated macroporous resins include pretreated SD300, D101, X-5, H103 and SV300.

[0012] Preferably, the emodin adsorbent solution is prepared as follows: crude emodin is dissolved in 25% ethanol at a concentration of 4.0-10.0 g / L, and the pH of the solution is adjusted to 9.0-12.0. While emodin is virtually insoluble in water, it is soluble in ethanol and alkaline solutions. However, high ethanol concentrations are not conducive to resin adsorption. Therefore, the present invention utilizes low-concentration ethanol and adjusts the pH to an alkaline level when preparing the emodin adsorbent solution. Crude emodin, in addition to the target compound emodin, also contains impurity peaks (such as chrysophanol and physophanol methyl ether). These impurities are organic compounds with similar structures to emodin, contributing to the low yield of currently available high-purity emodin and the difficulty of purification.

[0013] Preferably, the crude emodin is emodin with a purity of less than 95%.

[0014] Preferably, the adsorption time under static test conditions is 2.5 to 3.0 hours. During the static adsorption process, the adsorption capacity of emodin by the macroporous resin shows an overall upward trend as the adsorption time increases within the first 3 hours. After 3 hours, the adsorption capacity remains essentially constant. Therefore, under static test conditions, saturation is achieved within 2.5 to 3.0 hours. When the concentration of the emodin adsorbate solution increases to a certain level, impurity molecules compete with emodin for active sites in the macroporous resin, resulting in a decrease in the adsorption capacity of emodin. Therefore, the present invention has shown that the adsorption capacity of emodin by the macroporous resin is better when the concentration of the adsorbate solution is 7.5 to 8.0 g / L.

[0015] Preferably, the pretreated macroporous resin with the best adsorption effect is H103.

[0016] Preferably, the chromatographic analysis conditions are as follows: C18 chromatographic column (4.6 mm × 250 mm, 5 μm), mobile phase: methanol-0.1% phosphoric acid solution (82:18); flow rate: 1 mL·min -1 ; Detection wavelength: 220 nm; column temperature: 30 ℃; injection volume: 10 μL.

[0017] Preferably, the dynamic adsorption conditions are: a diameter-to-height ratio of 1:(3-7), a concentration of the emodin adsorbate solution of 7.5-8.0 g / L, and a sample loading flow rate of 0.3-1.0 mL / min. Under dynamic adsorption conditions, a higher resin height in the chromatography column reduces leakage of the target substance, increases the amount of target substance adsorbed by the resin, and allows for a relatively longer contact time between the resin and the sample solution. Most of the emodin is absorbed by the macroporous adsorption resin H103, resulting in a better dynamic adsorption effect. After experimental comparison, the optimal diameter-to-height ratio was determined to be 1:3. Due to intermolecular forces, emodin molecules are effectively adsorbed by the resin. As the sample loading flow rate slows and the sample concentration increases, the adsorption of emodin in the resin also increases. The optimal loading concentration is 8.0 g / L. In the present invention, the emodin adsorbate solution concentration during loading is set at 7.5-8.0 g / L. A slow loading flow rate is beneficial for increasing the resin's adsorption of emodin, with the optimal dynamic adsorption flow rate being 0.3 mL / min.

[0018] Preferably, desorption is performed using 25% to 75% ethanol at a flow rate of 0.3 to 0.5 mL / min. The emodin content in ethanol eluates of varying concentrations shows a trend of first increasing and then decreasing with increasing eluate volume. As the ethanol elution flow rate increases, the earlier the peak of the dynamic elution curve appears, the more the emodin content in the eluate is calculated. This indicates that a faster eluate flow rate results in a poorer desorption effect. The present invention selects 0.3 mL / min as the optimal dynamic desorption elution flow rate for the emodin solution.

[0019] The invention also provides high-purity emodin purified by the method.

[0020] Compared with the prior art, the present invention is beneficial in that:

[0021] 1. Compared with traditional purification of emodin, which relies more on trial and error and lacks theoretical support, the present invention uses molecular docking technology to analyze the interaction mechanism between emodin and macroporous resin at the microscopic level, and provides a scientific theoretical basis for process development through molecular dynamics simulation and binding energy calculation, avoiding blind screening and reducing trial and error time and cost.

[0022] 2. In terms of purification effectiveness, the present invention uses high-performance liquid chromatography to perform precise chromatographic analysis of samples before and after adsorption. By quantifying the differences in the main component and impurity peaks and separation, the method systematically screens for suitable macroporous resins. Static and dynamic adsorption and desorption are used to obtain eluents, and the eluents are concentrated and crystallized. The purity of emodin is increased to 99.8%. The process of the present invention is simple to operate, highly reproducible, and significantly reduces labor costs. The macroporous resin is recyclable, reducing consumables expenditures. It combines environmental and economic advantages, providing an efficient and stable technical solution for laboratory preparation and industrial production, and effectively promoting the high-quality development of the emodin industry. BRIEF DESCRIPTION OF THE DRAWINGS

[0023] Figure 1 The effect of different emodin adsorbent solutions on the equilibrium adsorption capacity;

[0024] Figure 2 is the static adsorption isotherm of emodin;

[0025] Figure 3 The pseudo-first-order kinetic simulation of the static adsorption of emodin;

[0026] Figure 4 It is a pseudo-second-order kinetic simulation of the static adsorption of emodin;

[0027] Figure 5 is the Weber-Morris intraparticle diffusion model;

[0028] Figure 6 N2 adsorption and desorption isotherms of macroporous resin H103 at 77 K;

[0029] Figure 7 is the pore size distribution of macroporous resin H103;

[0030] Figure 8 This is the molecular docking result of macroporous resin H103 and emodin;

[0031] Figure 9 HPLC chromatograms of crude emodin solution before (a) and after (b) purification by macroporous resin H103. DETAILED DESCRIPTION

[0032] The technical solution of the present invention will be described clearly and completely below. Obviously, the embodiments described are only some embodiments of the present invention, not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making creative work are within the scope of protection of the present invention. Equivalent transformations or substitutions of methods, process routes, and functions made by those skilled in the art based on the following embodiments are within the scope of protection of the present invention.

[0033] The present invention provides a process optimization method for purifying emodin with a macroporous resin, comprising the following steps:

[0034] Different pretreated macroporous resins were mixed with emodin adsorbate solutions and adsorbed under static test conditions until saturation. The solutions before and after adsorption were then chromatographically analyzed. Molecular docking, molecular dynamics, and binding energy were used to calculate the interaction mechanism between emodin and the macroporous resins, and the optimal macroporous resin and purification conditions were screened.

[0035] The macroporous resin with the best adsorption effect is used to adsorb the emodin adsorbate solution to saturation under dynamic adsorption conditions. The impurities are first washed with water and then desorbed with ethanol to obtain an eluate. The eluate is concentrated and crystallized to obtain high-purity emodin.

[0036] In some examples, the preparation method of the emodin adsorbent solution is as follows: dissolving crude emodin in 25% ethanol at a concentration of 4.0-10.0 g / L, and adjusting the pH of the solution to 9.0-12.0.

[0037] In some examples, the adsorption time was 2.5-3.0 h under static test conditions.

[0038] In some examples, the dynamic adsorption conditions are: diameter-to-height ratio of 1:(3-7), concentration of emodin adsorbate solution of 7.5-8.0 g / L, and flow rate of sample loading of 0.3-1.0 mL / min.

[0039] In some examples, desorption was performed with 25% to 75% ethanol at a flow rate of 0.3 to 0.5 mL / min.

[0040] The materials, reagents, and instruments used in the following examples are as follows:

[0041] Materials and reagents: Crude rhubarb was purchased from Jiuzhou Kangyuan Biotechnology; macroporous resins SD300, D101, X-5, H103, and SV300 were purchased from Zhejiang Zhengguang Industrial Co., Ltd.; ethanol, hydrochloric acid, and sodium hydroxide were purchased from Sinopharm Chemical Reagent Co., Ltd. All reagents used were of analytical grade unless otherwise specified.

[0042] Instruments and equipment: UV-2100 visible spectrophotometer, Unicol (Shanghai) Instrument Co., Ltd.; analytical balance and electrodes, Mettler, Switzerland; ultrasonic cleaner, Shenzhen Fangao Microelectronics Co., Ltd.; multifunctional constant temperature water bath oscillator, Shanghai Yiheng Instrument Co., Ltd.; SHZ-DIII circulating water multi-purpose vacuum pump, Yuhua Instrument Co., Ltd.; rotary evaporator, EYELA; glass chromatography column (Φ 20 mm × 180 mm), Changshu Aohua Instrument Co., Ltd.

[0043] Example 1

[0044] (1) Determination of emodin content

[0045] The absorbance value was measured at 530 nm by ultraviolet spectrophotometry and recorded. The standard curve was drawn with the content of emodin (mg / L) (C) as the horizontal axis and the absorbance value (A) as the vertical axis. The linear regression equation was obtained: A = 0.0034 × C + 0.014, R 2= 0.998, indicating that the content of emodin had a good linear relationship in the range of 0.001~0.045 g / L.

[0046] (2) Preparation of emodin adsorbent solution

[0047] The crude emodin was prepared into 8 g / L solutions using water, 5%, 25%, and 75% ethanol, respectively, and the pH was adjusted to 9.0, 10.0, 11.0, and 12.0 with alkali to obtain different emodin adsorbent solutions. 150 mL of the emodin adsorbent solution was placed in a stoppered conical flask, and 9 g of pretreated and drained H103 macroporous resin was weighed. The resin was subjected to constant temperature oscillation adsorption at a temperature of 293.15 K and a speed of 150 r / min for 8 h to allow for sufficient adsorption. The absorbance value was measured at 530 nm and substituted into the standard curve in step (1) to calculate the concentration. The equilibrium adsorption amount was calculated according to the following formula:

[0048]

[0049] Among them, q e is the equilibrium adsorption capacity, mg / g; V is the volume of the solution added in the experiment, mL; W is the weight of the dry resin, g; C0 is the initial concentration of the emodin adsorbate solution; C e is the concentration of the solution after full adsorption.

[0050] The equilibrium adsorption amount of emodin under various conditions is as follows Figure 1 As shown. Figure 1 It can be seen that the optimal preparation conditions for the emodin adsorbate solution are to use 25% volume fraction ethanol and adjust the pH to 11.0.

[0051] (3) Screening of the best resin

[0052] Depending on the nature of the substance being separated, the resin's adsorption strength varies. To comprehensively explore the adsorption performance of different resins for emodin, systematic chromatographic analysis of samples before and after adsorption was performed. By accurately measuring the peak values ​​and resolution of the main component and impurities, and comparing and analyzing the differences in the effects of different resin treatments, we provide a scientific basis for optimizing the separation and purification process of emodin.

[0053] The chromatographic analysis conditions were as follows: Tianjin Puxiang C18 column (4.6 mm × 250 mm, 5 μm), mobile phase: methanol-0.1% phosphoric acid solution (82:18); flow rate: 1 mL min -1 ; Detection wavelength: 220 nm; column temperature: 30℃; injection volume: 10 μL.

[0054] 9 g of pretreated and dried macroporous resins (SD300, D101, X-5, H103, and SV300) were weighed and 150 mL of 4, 8, and 10 g / L emodin adsorbent solutions (prepared with 25% ethanol by volume and adjusted to pH 11.0) were placed in stoppered Erlenmeyer flasks and placed in a thermostatic oscillator. The adsorption was carried out at a temperature of 293.15 K and a speed of 150 r / min for 8 h to allow for sufficient adsorption. The solutions before and after adsorption were analyzed by chromatography. The results are shown in Table 1. In Table 1, C0 is the initial concentration of the emodin adsorbent solution, A is the main component of emodin, B is an impurity with a similar structure to emodin, and C is the adsorbent. A0 is the initial concentration of the main component of emodin in the solution, C B0 is the initial concentration of impurities with similar structure to emodin in the solution, C Ae is the concentration of the main component of emodin in the solution after adsorption, C Be is the concentration of impurities with similar structures to emodin in the solution after adsorption, is the separation, q A is the equilibrium adsorption amount.

[0055] Table 1: Chromatographic analysis results before and after macroporous resin adsorption

[0056]

[0057] As shown in Table 1, the average adsorption capacity of H103 is the largest among the static equilibrium adsorption capacities of different types of macroporous resins for emodin adsorbate solution. After adsorption, the peak area of ​​the main component of emodin is significantly reduced, indicating that H103 macroporous resin has a strong adsorption effect on emodin and has no adsorption effect on impurities. The above results show that H103 macroporous resin has a better affinity for emodin than impurities, and the measured The separation value also shows that the use of H103 macroporous resin has the best separation effect between the main component of emodin and impurities.

[0058] (4) Adsorption thermodynamics test

[0059] Weigh 9 g of pretreated and dried H103 macroporous resin and place 150 mL of emodin adsorbent solutions with concentrations of 1, 4, 8, and 10 g / L (prepared with 25% ethanol and adjusted to pH 11.0) in stoppered Erlenmeyer flasks, respectively, in a constant temperature oscillator and adsorb at 150 r / min at temperatures of 283.15, 293.15, and 303.15 K for 8 h.

[0060] Static adsorption isotherm of emodin Figure 2 As shown by Figure 2It can be seen that with the increase of temperature, qe first increases and then decreases, indicating that appropriately increasing the temperature during the experiment is more conducive to adsorption. The adsorption amounts at temperatures of 303.15 K and 293.15 K are relatively close. Considering energy saving, it is most appropriate to set the adsorption temperature to 293.15 K.

[0061] Thermodynamic Freundlich, Dubinin-Radushkevich, and Temkin models revealed that the adsorption of emodin on H103 macroporous resin was preferential, with thermodynamic parameters ΔG < 0, ΔH > 0, and ΔS > 0. This indicates that the adsorption of emodin on H103 macroporous resin is spontaneous, easy, and has good affinity, demonstrating distinct physical adsorption characteristics.

[0062] (5) Adsorption kinetics test

[0063] Weigh 9 g of pretreated and dried H103 macroporous resin, place 150 mL of 8 g / L emodin adsorbent solution (prepared with 25% ethanol by volume and adjusted to pH 11.0) in a stoppered Erlenmeyer flask and place it in a constant temperature oscillator. Keep the temperature at 293.15 K and the oscillation speed at 150 r / min for 8 h. The results are as follows: Figures 3-5 shown.

[0064] Depend on Figures 3-5 The adsorption kinetics fitting curve showed that the adsorption equilibrium time of emodin on the macroporous adsorption resin H103 was 130 min, and the adsorption process was in accordance with the pseudo-first-order kinetic equation R 2 The value is greater than 0.99, and the adsorption rate is controlled by both intraparticle diffusion and membrane diffusion. In static tests, it was shown that H103 resin has the best adsorption performance for emodin.

[0065] (6) Molecular docking

[0066] In the above steps, the present invention screened macroporous resins SD300, D101, X-5, H103, and SV300, and the optimal macroporous resin was identified as H103. Macroporous resin H103 adsorbs the hydrophobic portion of the emodin molecule through weak intermolecular interactions. Furthermore, the macroporous resin H103 has a large specific surface area, which facilitates the flexible adsorption of emodin molecules through the pores. This was verified by a specific surface area test (BET) experiment. Based on various parameters, the nitrogen adsorption / desorption isotherm of macroporous resin H103 at 77 K and the pore size distribution diagram analyzed using the BJH model were obtained as follows: Figure 6 、 Figure 7As shown, its isotherm belongs to Type IV in the IUPAC classification. The adsorption process is monolayer to multilayer adsorption initially occurring on the mesopore walls, a common adsorption process in industry. Pores with diameters ranging from 2 to 50 nm are called mesopores, and pores with diameters greater than 50 nm are called macropores. The macroporous resin H103, calculated using the BJH model, consists primarily of 98% mesopores and 1.99% macropores, making it a mesoporous-macroporous adsorbent. The calculated average pore diameter is 13.8 nm. Calculations using Gaussian software indicate that the length of the emodin molecule is 1.37 Å. It is generally believed that adsorption is most favored when the pore diameter of the adsorbent is 2 to 6 times the molecular size of the adsorbate. The pore diameter of the macroporous resin H103 in the present invention is 100 times the diameter of the emodin molecule, making it highly favorable for emodin adsorption.

[0067] Molecular docking results of macroporous resin H103 and emodin (see Figure 8 ) indicates a binding energy of -4.0 kcal / mol. A binding energy less than 0 indicates that the emodin molecules spontaneously bind to the adsorbent. Subsequent three-dimensional interaction analysis revealed that two weak π-π bonds, with bond lengths of 2.9 and 2.64 Å, form between the main structure of the adsorbent H103 resin and the aromatic ring of the emodin molecule. Therefore, H103 resin is suitable for adsorbing emodin and exhibits optimal adsorption performance.

[0068] Example 2

[0069] (1) Preparation of emodin adsorbent solution: Crude emodin was dissolved in 25% ethanol and the pH was adjusted to 11.0 to obtain an emodin adsorbent solution with a mass concentration of 8 g / L.

[0070] (2) Soak the H103 macroporous adsorption resin in 98% ethanol for 24 h to remove the upper suspended particles, wash with distilled water until there is no alcohol smell, then wash with 1.5 mol / L hydrochloric acid, distilled water, 1.5 mol / L sodium hydroxide, and distilled water until neutral.

[0071] (3) The pretreated H103 macroporous adsorption resin was loaded into the chromatographic column by wet method, and the prepared emodin adsorbent solution was added thereto. Then, dynamic adsorption was adopted, wherein the adsorption conditions were: diameter-to-height ratio 1:7, sample flow rate 1 mL / min, and after adsorption, it was washed with distilled water until the effluent was colorless. The adsorption-saturated H103 macroporous adsorption resin was eluted with 150 mL of 25% by mass ethanol at a flow rate of 0.3 mL / min to obtain the eluate.

[0072] (4) The eluate is concentrated and crystallized to obtain the final high-purity product. The specific operation of concentration and crystallization is as follows: the eluate is cooled and adjusted to 20 °C, extracted and crystallized for 6 h until no more crystals are precipitated, filtered and dried to obtain the final emodin.

[0073] Example 3

[0074] (1) Preparation of emodin adsorbent solution: Crude emodin was dissolved in 25% ethanol and the pH was adjusted to 11.0 to obtain an emodin adsorbent solution with a mass concentration of 8 g / L.

[0075] (2) Soak the H103 macroporous adsorption resin in 98% ethanol for 24 h to remove the upper suspended particles, wash with distilled water until there is no alcohol smell, then wash with 1.5 mol / L hydrochloric acid, distilled water, 1.5 mol / L sodium hydroxide, and distilled water until neutral.

[0076] (3) The pretreated H103 macroporous adsorption resin was loaded into the chromatographic column by wet method, and the prepared emodin adsorbent solution was added thereto. Then, dynamic adsorption was adopted, wherein the adsorption conditions were: diameter-height ratio 1:5, sample flow rate 0.6 mL / min, and after adsorption, it was washed with distilled water until the effluent was colorless. The adsorption-saturated H103 macroporous adsorption resin was eluted with 130 mL of 50% by mass ethanol at a flow rate of 0.3 mL / min to obtain the eluate.

[0077] (4) The eluate is concentrated and crystallized to obtain the final high-purity product. The specific operation of concentration and crystallization is as follows: the eluate is cooled and adjusted to 20 °C, extracted and crystallized for 6 h until no more crystals are precipitated, filtered and dried to obtain the final emodin.

[0078] Example 4

[0079] (1) Preparation of emodin adsorbent solution: Crude emodin was dissolved in 25% ethanol and the pH was adjusted to 11.0 to obtain an emodin adsorbent solution with a mass concentration of 8 g / L.

[0080] (2) Soak the H103 macroporous adsorption resin in 98% ethanol for 24 h to remove the upper suspended particles, wash with distilled water until there is no alcohol smell, then wash with 1.5 mol / L hydrochloric acid, distilled water, 1.5 mol / L sodium hydroxide, and distilled water until neutral.

[0081] (3) The pretreated H103 macroporous adsorption resin was loaded into the chromatographic column by wet method, and the prepared emodin adsorbent solution was added thereto. Then, dynamic adsorption was adopted, wherein the adsorption conditions were: diameter-to-height ratio 1:3, sample flow rate 0.3 mL / min, and after adsorption, it was washed with distilled water until the effluent was colorless. The adsorption-saturated H103 macroporous adsorption resin was eluted with 120 mL of 75% by mass ethanol at a flow rate of 0.3 mL / min to obtain the eluate.

[0082] (4) The eluate is concentrated and crystallized to obtain the final high-purity product. The specific operation of concentration and crystallization is as follows: the eluate is cooled and the temperature is adjusted to 20°C, and then extracted and crystallized for 6 hours until no more crystals are precipitated. The product is then filtered and dried to obtain the final emodin.

[0083] Test Case

[0084] The purified samples were analyzed and verified by high-performance liquid chromatography (HPLC). Based on precise retention times and peak areas, Figure 9 shows that the impurity peaks with retention times between 0.8 and 1.1 min in the original sample have disappeared. A certain mass of crude emodin and macroporous resin-purified product were weighed, dissolved in 70% methanol, and the volume was adjusted to 10 mL. The purity was calculated according to the following formula. The purity results of the finished emodin products in each example are shown in Table 2.

[0085]

[0086] Wherein, P is the purity of emodin, %; M1 is the mass of emodin purified by macroporous resin, g; M0 is the mass of crude emodin, g.

[0087] Table 2: Purity results of emodin

[0088]

[0089] As shown in Table 2, the macroporous resin with the best adsorption effect was used to adsorb the emodin adsorbate solution to saturation under dynamic adsorption conditions. After washing with water, the eluate was desorbed with ethanol to obtain an eluate. The eluate was concentrated and crystallized to obtain high-purity emodin. The purity of emodin increased from 94.9% before purification to 99.8%, and the impurity removal rate was as high as 100%. This fully demonstrates that the H103 macroporous adsorption resin purification process of the present invention can effectively remove impurities, achieve efficient enrichment and purification of emodin, and provide a high-purity raw material guarantee for subsequent research and industrial production.

[0090] In summary, the present invention involves mixing different pretreated macroporous resins with an emodin adsorbate solution, subjecting the mixture to adsorption until saturation under static test conditions, performing chromatographic analysis on the solutions before and after adsorption, and using molecular docking, molecular dynamics, and binding energy to calculate the interaction mechanism between emodin and the macroporous resin to screen for the macroporous resin with the best adsorption effect and purification conditions. The macroporous resin with the best adsorption effect and the emodin adsorbate solution are then adsorbed to saturation under dynamic adsorption conditions, followed by washing with water and then desorption with ethanol to obtain an eluate. The eluate is then concentrated and crystallized to obtain high-purity emodin. The purified emodin has a purity of up to 99.8%, providing an efficient, simple, and stable method for obtaining high-purity emodin and for its industrial-scale production.

[0091] The above specific embodiments describe the implementation of the present invention in detail, but the present invention is not limited to the specific details of the above embodiments. Within the scope of the claims and technical concept of the present invention, various simple modifications and changes can be made to the technical solution of the present invention, and these simple modifications all fall within the scope of protection of the present invention.

Claims

1. A process optimization method for purifying emodin with macroporous resin, characterized in that: The following steps are involved: Different pretreated macroporous resins were mixed with emodin adsorbate solutions and adsorbed under static test conditions until saturation. The solutions before and after adsorption were then chromatographically analyzed. Molecular docking, molecular dynamics, and binding energy were used to calculate the interaction mechanism between emodin and the macroporous resins, and the optimal macroporous resin and purification conditions were screened. The macroporous resin with the best adsorption effect is used to adsorb the emodin adsorbate solution to saturation under dynamic adsorption conditions. The impurities are first washed with water and then desorbed with ethanol to obtain an eluate. The eluate is concentrated and crystallized to obtain high-purity emodin.

2. The process optimization method for purifying emodin with macroporous resin according to claim 1, wherein The preparation method of the emodin adsorbent solution is as follows: dissolving crude emodin in 25% ethanol at a concentration of 4.0-10.0 g / L, and adjusting the pH of the solution to 9.0-12.

0.

3. The process optimization method for purifying emodin with macroporous resin according to claim 2, characterized in that: The crude emodin has a purity of less than 95%.

4. The process optimization method for purifying emodin with macroporous resin according to claim 1, characterized in that: The adsorption time under static test conditions is 2.5~3.0 h.

5. The process optimization method for purifying emodin with macroporous resin according to claim 1, characterized in that: The macroporous resin with the best adsorption effect is H103.

6. The process optimization method for purifying emodin with macroporous resin according to claim 1, characterized in that: The method for pretreating the macroporous resin is as follows: soaking the macroporous resin in ethanol to remove upper suspended particles, and then sequentially washing with water, acid washing, water washing, alkali washing, and water washing to obtain the pretreated macroporous resin.

7. The process optimization method for purifying emodin with macroporous resin according to claim 1, characterized in that: The chromatographic analysis conditions were as follows: C18 column 4.6 mm × 250 mm, 5 μm; mobile phase: methanol-0.1% phosphoric acid solution; flow rate: 1 mL·min -1 Detection wavelength: 220 nm; Column temperature: 30°C; injection volume: 10 μL.

8. The process optimization method for purifying emodin with macroporous resin according to claim 1, characterized in that: The dynamic adsorption conditions are as follows: diameter-to-height ratio of 1:(3-7), concentration of emodin adsorbate solution of 7.5-8.0 g / L, and sample flow rate of 0.3-1.0 mL / min.

9. The process optimization method for purifying emodin with macroporous resin according to claim 1, characterized in that: Desorb with 25%~75% ethanol at a flow rate of 0.3~0.5mL / min.

10. High-purity emodin purified by the method according to any one of claims 1 to 9.

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