A template molecule of a multifunctional saponin compound molecularly imprinted polymer, the molecularly imprinted polymer and application thereof

Abstract

The present application relates to a kind of multifunctional saponins molecularly imprinted polymer template molecule, molecularly imprinted polymer and its application, belong to plant component refining technical field.The molecularly imprinted polymer of the present application uses virtual template molecule, the template molecule contains hydrogen bond donor and strong electronic effect, it has good adsorption performance to natural saponins compound molecules such as Jiumu saponin AIII, Jiumu saponin BII, Jiumu saponin BIII, Sanqi saponin R1, ginsenoside Rg1, ginsenoside Rg3, ginsenoside Rh2, Tribulus saponin K, Tribulus saponin D, Platanus orientalis saponin D, Shandeng saponin B, Chonglou saponin I, Chonglou saponin II, Chonglou saponin VI.The virtual template molecule is low in cost, can be used for the large-scale preparation of molecularly imprinted polymer, significantly reduces the extraction cost of natural saponins compound.

Description

TECHNICAL FIELD

[0001] The application belongs to the technical field of plant component refining, and particularly relates to a template molecule of a multifunctional saponin compound molecular imprinting polymer, a molecular imprinting polymer and application thereof. BACKGROUND

[0002] Molecular imprinting polymers are a new type of enrichment materials, which are widely used in the field of biochemical separation and analysis because of the stability of the material structure and the specificity in the recognition of target materials. By optimizing the template molecule, functional polymer unit and pore-forming agent system, the selectivity of the molecular imprinting polymer can be controlled, so that it has selective recognition ability with different widths to adapt to different application requirements. In the field of cosmetics, steroidal saponins, as a relatively safe, stable and effective skin care ingredient, have great application value. However, it is difficult to industrialize the application because it is difficult to separate a specific type of steroidal saponin from plants due to the similar structure of various steroidal saponins.

[0003] In summary, it has great research potential and commercial value to design and prepare a molecular imprinting polymer for enriching, purifying and analyzing specific types of steroidal saponins. SUMMARY

[0004] The first object of the present application is to provide a template molecule of a multifunctional saponin compound molecular imprinting polymer, which reduces the extraction cost by designing a virtual template molecule.

[0005] The second object of the present application is to provide a molecular imprinting polymer prepared from the above template molecule.

[0006] The third object of the present application is to provide an application of the above molecular imprinting polymer.

[0007] The object of the present application can be achieved by the following technical solutions.

[0008] A template molecule of a multifunctional saponin compound molecular imprinting polymer has the following structure:

[0009]

[0010] The molecular imprinting polymer prepared from the above template molecule has a preparation method comprising the following operations:

[0011] Preparation of the dispersed phase: mix the hydrophilic functional monomer, the crosslinking agent, water and the template molecule to prepare solution A;

[0012] Preparation of the continuous phase: mix the emulsifier with n-hexane under an inert gas atmosphere to form the continuous phase as solution B;

[0013] Preparation of the reverse phase dispersion system: under the atmosphere of inert gas, solution A is mixed with solution B to obtain a white emulsion;

[0014] Polymerization reaction: the white emulsion is mixed with an initiator to obtain a polymer microsphere mixture;

[0015] Post-treatment: the polymer microsphere mixture is mixed with methanol, the precipitate is obtained by filtration, the template molecules are eluted from the precipitate, and then the precipitate is washed to be neutral and dried to constant weight to obtain the molecularly imprinted polymer microspheres.

[0016] As a preferred scheme of the present application, in the preparation of the dispersion phase, the hydrophilic functional monomer is one of acrylamide, allylamine and methacrylic acid.

[0017] As a preferred scheme of the present application, in the preparation of the dispersion phase, the crosslinking agent is at least one of N, N'-methylenebisacrylamide, polyethylene glycol dimethacrylate and divinylbenzene.

[0018] As a preferred scheme of the present application, in the preparation of the dispersion phase, the molar ratio of the hydrophilic functional monomer and the template molecule is 1:4.5-8.5.

[0019] As a preferred scheme of the present application, in the preparation of the continuous phase, the emulsifier is at least one of docusate sodium and polyoxyethylene lauryl ether.

[0020] As a preferred scheme of the present application, in the polymerization reaction, the initiator is one of potassium persulfate, benzoyl peroxide and ammonium persulfate.

[0021] As a preferred scheme of the present application, in the polymerization reaction, the polymerization reaction temperature is 30-90℃.

[0022] As a preferred scheme of the present application, in the polymerization reaction, the polymerization reaction time is 1-4h, and the stirring is continuously performed during the polymerization reaction.

[0023] The above-mentioned molecularly imprinted polymer is applied to the separation of natural saponin compounds, and the natural saponin compounds are zhimu saponin AIII, zhimu saponin BII, zhimu saponin BIII, sanchi saponin R1, ginsenoside Rg1, ginsenoside Rg3, ginsenoside Rh2, tribuloside K, tribuloside D, platycodon saponin D, liriope saponin B, paris saponin I, paris saponin II and paris saponin VI.

[0024] The present application has the following beneficial effects:

[0025] The molecularly imprinted polymer of the present application uses (2R, 3R, 4S, 5R, 6R)-2-(hydroxymethyl)-6-(((2aR, 4S, 6aS, 6bS, 8aS, 8bR, 9S, 10R, 11aS, 12aS, 12bR)-5', 6a, 8a, 9-tetramethylicosahydrospiro[furo[2', 1': 4, 5]indeno[2, 1-b]furan-10, 2'-pyran]-4-yl)oxy)tetrahydro-2H-pyran-3, 4, 5-triol as a template molecule, which contains a hydrogen bond donor and a strong electronic effect, can react with 14 specific target products such as timosaponin AIII, timosaponin BII, timosaponin BIII, notoginseng saponin R1, ginsenoside Rg1, ginsenoside Rg3, ginsenoside Rh2, tribuloside K, tribuloside D, platycodon saponin D, liriope saponin B, paris saponin I, paris saponin II, paris saponin VI to form multiple action sites such as hydrogen bonds and hydrophobic interactions, etc. In the separation process, because the molecularly imprinted polymer can form stronger intermolecular forces and higher specific spatial pockets with specific types of saponin compounds, it has higher selectivity, and specific types of saponin compounds can be separated from natural products. In addition, the virtual template molecule is low in cost and can be used for large-scale preparation of molecularly imprinted polymers, significantly reducing the extraction cost of natural saponin compounds.

[0026] The natural saponin compound molecularly imprinted polymer prepared by the preparation method of the present application can increase the adsorption amount of the target molecule in the application process, improve the purity of the natural saponin compound, and can be used for the separation, enrichment and purification of natural saponin compounds in natural product extracts, has high enrichment degree, strong accuracy and wide application range, and has good adsorption performance for natural saponin compound molecules of timosaponin AIII, timosaponin BII, timosaponin BIII, notoginseng saponin R1, ginsenoside Rg1, ginsenoside Rg3, ginsenoside Rh2, tribuloside K, tribuloside D, platycodon saponin D, liriope saponin B, paris saponin I, paris saponin II, and paris saponin VI. DETAILED DESCRIPTION

[0027] The present application will be further described in detail below in combination with the specific embodiments.

[0028] Example 1

[0029] A template molecule (2R,3R,4S,5R,6R)-2-(hydroxymethyl)-6-(((2aR,4S,6aS,6bS,8aS,8bR,9S,10R,11aS,12aS,12bR)-5',6a,8a,9-tetramethylicosahydrospiro[furo[2',1':4,5]indeno[2,1-b]furan-10,2'-pyran]-4-yl)oxy)tetrahydro-2H-pyran-3,4,5-triol, having the structural formula:

[0030]

[0031] The specific experimental operations are as follows:

[0032] (1) In a round-bottom flask, 10.0 mmol of D-glucose and 10 mL of acetic anhydride were added to obtain a mixture, and the mixture was cooled to 0°C. Then 0.1 m of perchloric acid (70 wt%) was added dropwise to the mixture while stirring, and after the dropwise addition was completed, the stirring was continued at 0°C for 5 h to obtain a reaction solution. The reaction solution was diluted with dichloromethane (40 mL). The diluted reaction solution was washed with water (20 mL), saturated NaHCO3 solution (20 mL), and brine (20 mL), respectively. The organic layer was dried with anhydrous sodium sulfate, and then the organic solvent in the organic layer was removed by a rotary evaporator to obtain an acetyl-protected glucose, with a yield of 96%.

[0033] (2) In a round-bottom flask, 4.0 mmol of acetyl-protected glucose of compound A and 40 mL of dichloromethane were added to obtain a mixed solution, and 1.66 mL of 48% mass fraction of boron trifluoride ether solution (item number: Titan Technology 01375737) was added dropwise to the mixed solution at 0°C. After the mixture was continuously stirred at 0°C for 10 min, 5.2 mmol of compound B (Sarsasapogenin; CAS number: 82597-74-8) was added to the mixture, and then the temperature was raised to room temperature and the stirring was continued for 24 h. After the reaction was completed, 20 mL of saturated NaHCO3 aqueous solution was added to the mixture at 0°C, and the organic layer was washed with water (20 mL x 3). After separation, the water layer was further extracted with 40 mL of dichloromethane. The combined dichloromethane organic phase was dried with anhydrous sodium sulfate and concentrated under reduced pressure, and compound C was obtained after column chromatography purification, with a yield of 85%.

[0034] (3) 3.0 mmol of compound C was dissolved in a mixture of 20 mL of methanol and 3.0 mL of 25% mass fraction of aqueous ammonia solution, and after stirring at room temperature overnight, the solvent was removed under reduced pressure, and the template molecule was obtained after column chromatography purification, with a yield of 91%.

[0035] The template molecule has the following spectral data: 1H NMR (400 MHz, CDC13) δ 5.50 - 5.39 (m, 1H), 4.78, (s, 1H), 4.68, (s, 2H), 4.55 (m, 1H), 4.40 (m, 1H), 4.10 (m, 1H), 3.95 (dd, J = 11.0, 2.7 Hz, 1H), 3.65 (m, 6H), 3.30 (d, J = 11.0 Hz, 1H), 2.28 (m, 2H), 2.12 (m, 2H), 2.09 - 1.95 (m, 5H), 1.88 (m, 2H), 1.85 - 1.65 (m, 6H), 1.60 - 1.35 (m, 4H), 1.33 - 1.27 (m, 2H), 1.26 - 1.09, (m, 4H), 1.08 - 1.02, (d, J = 7.1 Hz, 3H), 1.02 - 0.98 (d, J = 6.9 Hz, 3H), 0.97 (s, 3H), 0.76 (s, 3H). 13 C-NMR (100 MHz, CDC13,) δ 118.9, 109.7, 81.8, 81.0, 76.6, 74.3, 71.6, 67.1, 65.1, 62.3, 62.1, 56.5, 42.1, 40.3, 39.8 (2C), 36.5, 35.3 (2C), 33.5 (2C), 29.9, 27.8 (2C), 27.1 (2C), 26.5 (2C), 23.9, 20.8, 16.5 (2C), 14.3. HRMS (ESI) m / z calcd for C 33 H 54 O8[M+H] + : 579.3824, found: 579.4028.

[0036] Preparation and performance test of molecularly imprinted polymer of example 2

[0037] 1. Preparation of molecularly imprinted polymer, the steps are as follows:

[0038] 1.1 Preparation of the dispersed phase: 6.5 mmol of methacrylic acid and 32.5 mmol of N,N'-methylenebisacrylamide (MBA) were dissolved in 35 mL of water, and 1 mmol of the template molecule (2R,3R,4S,5R,6R)-2-(hydroxymethyl)-6-(((2aR,4S,6aS,6bS,8aS,8bR,9S,10R,11aS,12aS,12bR)-5',6a,8a,9-tetramethylicosahydrospiro[furo[2',1':4,5]indeno[2,1-b]furan-10,2'-pyran]-4-yl)oxy)tetrahydro-2H-pyran-3,4,5-triol prepared in Example 1 were added and uniformly dispersed, and the pre-polymer was formed by magnetic stirring at room temperature for 0.5 h to obtain solution A;

[0039] 1.2 Preparation of the continuous phase: The continuous phase was prepared by dissolving 13 g of sodium docusate (AOT) and 25 g of L4 polyoxyethylene lauryl ether (BRIJ 30) in 500 mL of n-hexane under a nitrogen atmosphere to form solution B;

[0040] 1.3 Preparation of the reverse-phase dispersion system: Solution A was slowly transferred to solution B while stirring, and stirring was continued at room temperature under a nitrogen atmosphere for 60 min to obtain a white emulsion;

[0041] 1.4 Polymerization reaction: 0.8 mL of 10% (w / v) ammonium persulfate (APS) was added to the white emulsion to initiate the polymerization reaction, and the reaction was continued at 50°C with stirring at 320 r / min for 3 h to obtain a polymer microsphere mixture;

[0042] 1.5 Post-treatment: Methanol was added to the polymer microsphere mixture to cause the polymer to aggregate and precipitate, and the polymer was filtered to obtain a polymer. The dried polymer was placed in a Soxhlet extractor, and the template molecule in the polymer was extracted with ethanol / ice acetic acid (9:1, V / V). The polymer was dried to a constant weight at 55°C under vacuum, and the imprinted material was classified using different mesh screens after drying to obtain molecularly imprinted polymers of different sizes.

[0043] 2. Performance test

[0044] 2.1 Study on the adsorption performance for ginsenoside Rg3:

[0045] i) Total saponins detection method (for simplifying the characterization of ginsenoside Rg3): Prepare 500 μg / mL oleanolic acid standard solution with methanol as solvent, accurately pipette 40, 60, 100, 120, 200 μL of the control sample solution into 10 mL test tubes with stoppers, and place them in a boiling water bath to evaporate the solvent, then add 0.2 mL of freshly prepared 5 wt% vanillin-glacial acetic acid solution, shake well, then add 0.8 mL of perchloric acid, shake well, then place them in a 70°C constant temperature water bath for 30 minutes, immediately cool with ice water for 5 minutes, add glacial acetic acid to 10 mL, shake well; use the reagent as a blank. After eliminating bubbles, determine the absorbance at a wavelength of 546 nm using a UV spectrophotometer, and plot a standard curve with absorbance as the vertical coordinate and concentration as the horizontal coordinate. Dilute the sample to be tested with methanol to a concentration of 20-100 μg / mL, and take 1 mL for detection, and calculate the content by the standard curve.

[0046] ii) Determination of adsorption capacity: accurately weigh 0.2 g of molecularly imprinted polymer into a ground conical flask, add 5.0 mL of 3 mg / mL ginsenoside Rg3 methanol solution, and constant temperature oscillation adsorption for 5 h, then centrifugal separation, dilute the upper clear solution with methanol to a concentration of 20-100 μg / mL, then determine the absorbance 3 times in parallel according to method i, take the arithmetic mean, and calculate the total saponin concentration by the standard curve. The adsorption capacity of the imprinted polymer is calculated from the change in total saponin concentration in the solution before and after adsorption (60.7 mg / g). The calculation formula is:

[0047] Q = (C0-C)*V / W

[0048] Wherein: Q: adsorption capacity per gram of polymer (mg / g);

[0049] C0: original mass concentration of the adsorbed compound in the equilibrium adsorption experiment (mg / L);

[0050] C: mass concentration of the adsorbed compound in the upper clear solution after reaching adsorption equilibrium (mg / L);

[0051] V: volume of the adsorption solution (L);

[0052] W: mass of the imprinted polymer (g).

[0053] Test Example 1: Investigation of the effect of different emulsifiers on the shape of the polymer

[0054] Based on the preparation method in Example 2, different emulsifiers were used as dispersants (see Table 1) to obtain molecularly imprinted polymers. The adsorption capacity of each polymer for ginsenoside Rg3 was determined, and the results are shown in Table 1:

[0055] Table 1

[0056] Emulsifier Polymer adsorption amount Sodium docusate (AOT) 59.43 mg / g L4 Polyoxyethylene lauryl ether (BRIJ 30) 52.90 mg / g Triton X 100 24.05 mg / g

[0057] Table 1 shows that molecularly imprinted polymers prepared using sodium docusate (AOT) and L4 polyoxyethylene lauryl ether (BRIJ 30) as emulsifiers exhibited strong adsorption capacity for ginsenoside Rg3. The molecularly imprinted polymer prepared using Triton X100 as an emulsifier showed slightly lower adsorption capacity for ginsenoside Rg3 compared to the other two.

[0058] Experimental Example 2 investigated the effect of different types and proportions of functional monomers in the dispersed phase preparation step on the properties of the molecularly imprinted polymer.

[0059] Based on the preparation method in Example 2, different types and proportions of functional monomers were used to obtain different molecularly imprinted polymers, and their adsorption capacity for ginsenoside Rg3 was measured. The results are shown in Table 2.

[0060] Table 2

[0061] Table 2 shows that when acrylamide, allylamine, and methacrylic acid are used as hydrophilic functional monomers, and the template molecule is used in combination with them at a molar ratio of 1:6.5, the resulting molecularly imprinted polymers all exhibit good adsorption capacity for ginsenoside Rg3. The adsorption capacity is highest (61.24 mg / g) when the template molecule is used with methacrylic acid at a molar ratio of 1:6.5. The adsorption capacity of the molecularly imprinted polymers for ginsenoside Rg3 varies significantly when the template molecule and acrylamide are used in different molar ratios. The adsorption capacity decreases significantly (16.70 mg / g) when the molar ratio is 1:2.5, and is slightly lower than that when the molar ratio is 1:4.5 and 1:8.5.

[0062] Experimental Example 3 investigated the effect of using different crosslinking agents in the dispersed phase preparation step on the molecularly imprinted polymer in the preparation method.

[0063] Based on the preparation method in Example 2, different molecularly imprinted polymers were obtained by using different crosslinking agents (see Table 3). The changes in their molding rate, size, etc. were observed, and the results are shown in Table 3.

[0064] Table 3

[0065] Crosslinking agent Polymer adsorption amount N, N'-Methylene bisacrylamide (MBA) 60.39 mg / g Polyethylene glycol dimethacrylate (EGDMA) 54.71 mg / g Divinylbenzene (DVB) 30.09 mg / g

[0066] Table 3 shows that N,N'-methylenediacrylamide and polyethylene glycol dimethacrylate were used as crosslinking agents to prepare different molecularly imprinted polymers, all of which showed good adsorption of ginsenoside Rg3. The adsorption effect of using divinylbenzene was slightly lower than that of the other two.

[0067] Experimental Example 4 investigated the effect of using different initiators in the polymerization reaction step on the properties of the molecularly imprinted polymer in the preparation method.

[0068] Based on the preparation method in Example 2, different initiators were used (see Table 4) to observe the reaction performance of the different molecularly imprinted polymers. The results are shown in Table 4.

[0069] Table 4

[0070] Initiator Polymerization reaction Potassium persulfate Simple operation, rapid reaction, stable process Benzoyl peroxide Simple operation, rapid reaction, slightly poor process stability Ammonium persulfate Simple operation, rapid reaction, stable process

[0071] Table 4 shows that different molecularly imprinted polymers were prepared by using potassium persulfate, benzoyl peroxide, and ammonium persulfate as initiators. All of them were simple to operate and reacted rapidly. The difference was that the molecularly imprinted polymers prepared by potassium persulfate and ammonium persulfate had a stable reaction process, while the molecularly imprinted polymer prepared by benzoyl peroxide had a slightly less stable reaction process.

[0072] Experimental Example 5 investigated the effect of different reaction conditions in the polymerization step on the properties of the molecularly imprinted polymer in the preparation method.

[0073] Based on the preparation method in Example 2, different molecularly imprinted polymers were prepared by changing the reaction conditions of the polymerization reaction, including reaction temperature, reaction time, and mechanical stirring speed (see Table 5). The adsorption capacity of each polymer for ginsenoside Rg3 was measured, and the results are shown in Table 5.

[0074] Table 5

[0075]

[0076] Table 5 shows that different molecularly imprinted polymers prepared under polymerization conditions of 30–90℃ and 1–4 h can all form microspheres. The molecularly imprinted polymers prepared at 50℃ for 3 h and 70℃ for 2 h both exhibit good adsorption capacity for ginsenoside Rg3. However, the adsorption capacity of the molecularly imprinted polymer microspheres prepared at 30℃ for 4 h and 90℃ for 1 h is slightly lower for ginsenoside Rg3.

[0077] Example 3: Study on the adsorption performance of the molecularly imprinted polymer obtained in Example 2 for various natural saponins:

[0078] The adsorption performance test of ginsenoside Rg3 was conducted according to the method in 2.1, except that the test molecule was replaced with a substitute molecule (natural saponin compound molecule). The data obtained are shown in Table 6.

[0079] Table 6

[0080] Number Chemical name CAS number Adsorption amount (mg / g) 1 Timosaponin AIII 41059-79-4 52.70 2 Timosaponin BII 136656-07-0 56.34 3 Timosaponin BIII 142759-74-8 49.55 4 Panax notoginseng saponin R1 80418-24-2 52.41 5 Ginsenoside Rg1 22427-39-0 53.10 6 Ginsenoside Rh2 78214-33-2 57.27 7 Tragacanth saponin K 193605-07-1 52.54 8 Tragacanth saponin D 179464-23-4 57.66 9 Platycodon saponin D 58479-68-8 49.05 10 Liriope saponin B 182284-68-0 52.39 11 Chonglou saponin I 50773-41-6 52.96 12 Chonglou saponin II 76296-72-5 51.75 13 Chonglou saponin VI 55916-51-3 53.22

[0081] As shown in Table 6, the molecularly imprinted polymer prepared in Example 2 exhibits good adsorption properties for natural saponin compounds such as Anemarrhena asphodeloides AIII, Anemarrhena asphodeloides BII, Anemarrhena asphodeloides BIII, Panax notoginseng saponin R1, Ginsenoside Rg1, Ginsenoside Rh2, Tribulus terrestris saponin K, Tribulus terrestris saponin D, Platycodon grandiflorus saponin D, Ophiopogon japonicus saponin B, Paris polyphylla saponin I, Paris polyphylla saponin II, and Paris polyphylla saponin VI.

[0082] Example 4 tests the application of the molecularly imprinted polymer obtained in Example 2.

[0083] To verify the separation performance and application potential of this molecularly imprinted polymer, it was used as an adsorbent material for internal phase extraction to enrich and separate anemarrhena saponin BII from the extract of Anemarrhena asphodeloides.

[0084] The specific experiment is as follows:

[0085] (1) Preparation of Anemarrhena asphodeloides extract: Weigh 0.5g of Anemarrhena asphodeloides powder, add 80mL of 60% ethanol solution, and extract by ultrasonic reflux for 0.5h. After centrifugation, separate the supernatant and repeat the extraction of residue 3 times. Combine the supernatants, filter with a 0.1-micron membrane, and collect the filtrate. Concentrate the filtrate by rotary evaporator to obtain Anemarrhena asphodeloides extract (medicinal material: extract = 5:1, W / W). Seal and keep at low temperature for later use.

[0086] (2) Adsorption experiment: First, 200 mg of molecularly imprinted polymer was loaded into a solid-phase extraction column and activated with 5 mL of methanol. Then, a 20 mg / mL sample loading solution of Anemarrhena asphodeloides extract was prepared with methanol, and 3 mL of the sample loading solution was loaded onto the column. The column was then rinsed with 4 mL of methanol and eluted with ethanol-acetic acid (95:5, v / v).

[0087] (3) Results analysis: The collection solution was tested by high performance liquid chromatography according to the detection method of Anemarrhena saponin BII in Anemarrhena asphodeloides in Chinese Pharmacopoeia 2015 P213. The concentration of Anemarrhena saponin BII in the solution before and after adsorption was calculated. The recovery rate was 82.4%.

[0088] The above description is merely an embodiment of the present invention and does not limit the scope of patent protection. Any non-substantial changes or substitutions made by those skilled in the art based on the present invention will still fall within the scope of patent protection.

Claims

1. A method for separating and extracting natural saponin compounds, characterized in that, Includes the following steps: Step A. Prepare a molecularly imprinted polymer using template molecules. Step B. Use the molecularly imprinted polymer to separate and extract natural saponin compounds; The template molecular structure described in step A is as follows: Step A includes the following operations: Preparation of the dispersed phase: Mix the hydrophilic functional monomer, crosslinking agent, water and template molecules to prepare solution A; Preparation of continuous phase: The emulsifier is mixed with n-hexane under an inert gas atmosphere to form a continuous phase, which is solution B; the emulsifier is at least one of sodium docusate and polyoxyethylene lauryl ether. Preparation of a reversed-phase dispersion system: Solution A and solution B were mixed under an inert gas atmosphere to obtain a white emulsion; Polymerization reaction: The white emulsion is mixed with an initiator to obtain a polymer microsphere mixture; Post-processing: The polymer microsphere mixture was mixed with methanol, filtered to obtain the precipitate, the template molecules were washed off the precipitate until it was neutral, and then dried to constant weight to obtain molecularly imprinted polymer microspheres. The natural saponin compounds are one or more of the following: Anemarrhena saponin AIII, Anemarrhena saponin BII, Anemarrhena saponin BIII, Panax notoginseng saponin R1, Ginsenoside Rg1, Ginsenoside Rg3, Ginsenoside Rh2, Tribulus saponin K, Tribulus saponin D, Platycodon saponin D, Ophiopogon saponin B, Paris saponin I, Paris saponin II and Paris saponin VI.

2. The method for separating and extracting natural saponin compounds according to claim 1, characterized in that, In the preparation of the dispersed phase, the hydrophilic functional monomer is one of acrylamide, allylamine, and methacrylic acid.

3. The method for separating and extracting natural saponin compounds according to claim 1, characterized in that, In the preparation of the dispersed phase, the crosslinking agent is at least one of N,N'-methylenediacrylamide, polyethylene glycol dimethacrylate, and divinylbenzene.

4. The method for separating and extracting natural saponin compounds according to claim 1, characterized in that, In the preparation of the dispersed phase, the molar ratio of the hydrophilic functional monomer to the template molecule is 1:4.5 to 8.

5.

5. The method for separating and extracting natural saponin compounds according to claim 1, characterized in that, In the polymerization reaction, the initiator is one of potassium persulfate, benzoyl peroxide, and ammonium persulfate.

6. The method for separating and extracting natural saponin compounds according to claim 1, characterized in that, In the polymerization reaction, the polymerization temperature is 30–90℃.

7. The method for separating and extracting natural saponin compounds according to claim 1, characterized in that, In the polymerization reaction, the polymerization time is 1 to 4 hours, and stirring is carried out continuously during the polymerization process.

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