A milky-banana extract having acetylcholinesterase inhibitory activity and a preparation method thereof
By employing a combination of enzymatic hydrolysis, dual-frequency ultrasound, and macroporous resin combined with phospholipid complex technology, the problems of weak acetylcholinesterase inhibitory activity and difficulty in crossing the blood-brain barrier in jellyfish snow lotus extract have been solved, achieving a highly effective treatment for Alzheimer's disease.
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- QINGHAI UNIV FOR NATITIES
- Filing Date
- 2026-04-29
- Publication Date
- 2026-06-05
AI Technical Summary
Existing methods for extracting Saussurea medusa have failed to effectively break down cell walls, resulting in low dissolution rates and severe co-extraction of impurities. This leads to weak acetylcholinesterase inhibitory activity, and the extracts have difficulty crossing the blood-brain barrier, resulting in low bioavailability and limiting their efficacy in treating Alzheimer's disease.
An integrated approach combining enzymatic hydrolysis to disrupt cell walls, dual-frequency ultrasound-assisted extraction, and macroporous resin directional elution, along with phospholipid complex technology, was employed to target and enrich the acetylcholinesterase-inhibiting active components in Saussurea medusa. Furthermore, the phospholipid complex was used to improve the permeability of the blood-brain barrier.
It significantly increased the content and bioavailability of acetylcholinesterase inhibitory active ingredients in jellyfish snow lotus extract, enhanced its distribution in the central nervous system, improved learning and memory dysfunction, and had lower toxicity than chemically synthesized drugs.
Abstract
Description
Technical Field
[0001] This invention relates to the field of traditional Chinese medicine extraction and neuropharmaceutical technology, specifically to a jellyfish snow lotus extract with acetylcholinesterase inhibitory activity and its preparation method. Background Technology
[0002] Alzheimer's disease (AD) is a neurodegenerative disease of the central nervous system characterized by progressive cognitive impairment and memory loss. It has become the fourth leading cause of death among the elderly, after cardiovascular disease, cancer, and stroke. One of the main pathological features of AD is a significant decrease in the level of acetylcholine (ACh) in the cerebral cortex and hippocampus, leading to cholinergic neurotransmission dysfunction and consequently, a decline in learning and memory abilities.
[0003] Acetylcholinesterase (AChE) is a key enzyme in the body that rapidly hydrolyzes the neurotransmitter acetylcholine. Inhibiting AChE activity reduces the breakdown of acetylcholine, increases the concentration of acetylcholine in the synaptic cleft, and thus improves cholinergic neurotransmission. This is currently the most mature and widely used drug strategy for treating Alzheimer's disease (AD). First-line drugs such as donepezil and rivastigmine are acetylcholinesterase inhibitors. However, these chemically synthesized drugs generally have short half-lives, require long-term and frequent administration, have significant hepatotoxicity and nephrotoxicity, cause significant gastrointestinal reactions, and experience rapid symptom rebound after discontinuation, severely limiting their clinical application.
[0004] Saussurea medusa, a perennial herb belonging to the genus Saussurea in the family Asteraceae, is mainly distributed in alpine scree slopes and near the snow line at altitudes above 3000 meters on the Qinghai-Tibet Plateau. It is an important medicinal plant in the Tibetan medicine system. Traditional Tibetan medicine believes that Saussurea medusa has the effects of clearing heat and detoxifying, dispelling wind and dampness, promoting blood circulation, anti-inflammatory and analgesic effects, and anti-aging effects. It is often used to treat rheumatoid arthritis, menstrual disorders, altitude sickness, and other diseases. Modern research shows that Saussurea medusa is rich in flavonoids, phenolic acids, terpenes, volatile oils, and other active ingredients, possessing pharmacological effects such as antioxidant, anti-inflammatory, and immunomodulatory properties.
[0005] Traditional and existing research has focused on its anti-rheumatism, anti-aging, and gynecological menstrual regulation. Chemical composition research mainly revolves around total flavonoids and chlorogenic acid components. However, there is very little research on systematic screening of its acetylcholinesterase inhibitory activity, targeted enrichment of active ingredients, and anti-Alzheimer's disease applications. Its medicinal value has not been fully developed.
[0006] Current methods for extracting Saussurea medusa extract primarily employ simple water decoction, single-solvent reflux extraction, or conventional ultrasonic extraction. These methods fail to target the physicochemical properties of AChE-inhibiting active components (mainly small-molecule phenolic acids and specific polar flavonoid glycosides). Traditional extraction methods suffer from problems such as insufficient cell wall disruption, low dissolution rates of active components, and severe co-extraction of impurities (e.g., polysaccharides, proteins, chlorophyll), resulting in low levels of target active components, weak AChE inhibitory activity, and significant challenges and costs associated with subsequent purification.
[0007] Common Chinese herbal extracts are mostly multi-component mixtures with high polarity, making it difficult for them to effectively cross the blood-brain barrier (BBB). The BBB is a selective permeability barrier composed of brain capillary endothelial cells, the basement membrane, pericytes, and astrocyte foot processes, restricting most polar molecules and drugs from entering the central nervous system. Common jellyfish and snow lotus extracts have low oral bioavailability and insufficient concentrations in the central nervous system, severely limiting their efficacy against Alzheimer's disease.
[0008] Therefore, this application proposes a jellyfish snow lotus extract with acetylcholinesterase inhibitory activity and its preparation method. Summary of the Invention
[0009] The purpose of this invention is to provide a jellyfish snow lotus extract with acetylcholinesterase inhibitory activity and its preparation method, so as to solve the problems mentioned in the background art.
[0010] To achieve the above objectives, the present invention provides the following technical solution:
[0011] In a first aspect, a method for preparing a jellyfish snow lotus extract with acetylcholinesterase inhibitory activity, the preparation method comprising the following steps:
[0012] Step A: Complex enzymatic hydrolysis to break cell walls
[0013] Take the dried whole plant of Saussurea medusa, pulverize and sieve to obtain Saussurea medusa coarse powder. Add an acetate-sodium acetate buffer solution with a pH of 4.5-5.0 to the coarse powder, with the amount of buffer added being 1:10-1:20 (w / v) as the material-to-liquid ratio. Add a compound enzyme to the above mixture, the compound enzyme consisting of cellulase and pectinase in a mass ratio of 1.5-2.5:1, the total amount of the compound enzyme being 0.8%-1.5% of the mass of the coarse powder. Place the above mixture in a constant temperature water bath at 40-50℃ and enzymatically hydrolyze for 1.5-2.5 hours under shaking conditions to destroy the plant cell walls and intercellular layers, releasing the internal active substances, and obtaining the enzymatic hydrolysate.
[0014] Step B, Dual-frequency ultrasound-assisted extraction
[0015] The enzymatic hydrolysate obtained in step A was placed in an ultrasonic extraction device and extracted using a dual-frequency alternating ultrasonic method. The dual-frequency alternating ultrasonic method alternated between 40kHz and 100kHz frequencies, switching frequencies every 8-12 minutes. The ultrasonic power was 200W-400W, and the extraction time was 25-35 minutes. The cavitation effect and mechanical vibration generated by the dual-frequency ultrasound effectively disrupted organelle membrane structures, promoted the dissolution of small molecule phenolic acids and flavonoid glycosides, and improved extraction efficiency.
[0016] Step C: Dynamic adsorption and directional elution of macroporous resin
[0017] The extract obtained in step B is subjected to solid-liquid separation, and the supernatant is collected. The supernatant is then loaded onto a chromatography column packed with HPD-100 macroporous adsorption resin at a flow rate of 1.5-2.5 BV / h (bed volume / hour) for dynamic adsorption. After adsorption, elution is first performed with 4-6 BV of deionized water or distilled water at a flow rate of 2-3 BV / h to remove water-soluble impurities such as polysaccharides, proteins, and inorganic salts. Subsequently, a 60%-70% (v / v) aqueous ethanol solution is used as the eluent for directional elution at a flow rate of 1.0-2.0 BV / h, and the ethanol eluent is collected.
[0018] Step D, Concentration and Drying
[0019] The ethanol eluent collected in step C was concentrated under vacuum thin-film treatment at 45℃-55℃ and a vacuum degree of -0.06-0.09 MPa until the relative density reached 1.05-1.15 (measured at 60℃), yielding a concentrated solution. The concentrated solution was then spray-dried, with the inlet air temperature controlled at 170℃-190℃ and the outlet air temperature at 80-90℃, to obtain a pale yellow to yellowish-brown powder extract.
[0020] Preferably, in step A, the pulverization and sieving are performed through a 40-80 mesh sieve, the pH value is 4.8, the material-to-liquid ratio is 1:15, the mass ratio of cellulase to pectinase is 2:1, the total enzyme content is 1.2%, the enzymatic hydrolysis temperature is 45℃, and the enzymatic hydrolysis time is 2 hours.
[0021] Preferably, in step B, the dual-frequency alternating ultrasound switches frequencies every 10 minutes, the ultrasound power is 300W, and the extraction time is 30 minutes.
[0022] Preferably, in step C, the sample loading flow rate is 2 BV / h, the deionized water elution volume is 5 BV and the flow rate is 2.5 BV / h, and the volume fraction of the ethanol aqueous solution is 65% with an elution flow rate of 1.5 BV / h.
[0023] Preferably, in step D, the concentration temperature is 50°C, the vacuum degree is -0.08 MPa, and the relative density is 1.10; the inlet air temperature of the spray drying is 180°C, and the outlet air temperature is 85°C.
[0024] Secondly, a jellyfish snow lotus extract
[0025] The extract is prepared by the preparation method described in the first aspect above.
[0026] The quality control of the extract was performed using high-performance liquid chromatography (HPLC) fingerprinting. The extract contains chlorogenic acid, 1,5-dicaffeoylquinic acid, and luteolin-7-O-glucoside, with the total mass fraction of these three main active ingredients not less than 35%. Among them, the mass fraction of chlorogenic acid compounds (including chlorogenic acid, 1,5-dicaffeoylquinic acid, and other caffeoylquinic acid derivatives) is not less than 15%, and the mass fraction of flavonoid glycosides (including luteolin-7-O-glucoside and other flavonoid glycosides) is not less than 10%.
[0027] The half-maximal inhibitory concentration (IC50) of the extract against acetylcholinesterase in electric eels was determined by in vitro enzyme activity inhibition assay. 50 The value is 15-22 ug / mL, preferably 18.5 ug / mL.
[0028] Thirdly, a pharmaceutical composition
[0029] The pharmaceutical composition comprises the jellyfish snow lotus extract described in technical solution 2 above and a pharmaceutically acceptable carrier, wherein the carrier comprises a phospholipid material, and the extract and the phospholipid material form a phospholipid complex.
[0030] Specifically, the extract and soybean lecithin are dissolved in anhydrous ethanol at a mass ratio of 1:(1.5~2.5), and stirred at 35℃~45℃. The organic solvent is then removed by rotary evaporation to form a solid dispersion, i.e., a phospholipid complex. The phospholipid complex can be further mixed with pharmaceutically acceptable excipients to prepare oral solid dosage forms such as capsules, tablets, granules, or pills.
[0031] Preferably, the mass ratio of the extract to soybean lecithin is 1:2; the dissolution temperature is 40°C; and the phospholipid is one or more of soybean lecithin, egg yolk lecithin, hydrogenated soybean lecithin, or phosphatidylcholine.
[0032] Fourthly, the use of the jellyfish and snow lotus extract described in technical solution two or the pharmaceutical composition described in technical solution three in the preparation of drugs for the prevention and / or treatment of Alzheimer's disease.
[0033] The drug exerts its anti-Alzheimer's effect by inhibiting acetylcholinesterase activity, increasing acetylcholine levels in the brain, and improving learning and memory impairment.
[0034] This invention integrates "compound enzymatic hydrolysis for cell wall disruption, dual-frequency ultrasound-assisted extraction, and macroporous resin-guided elution" to target and enrich the AChE-inhibiting active components (small molecule phenolic acids and flavonoid glycosides) in Saussurea medusa by targeting their physicochemical properties. Compound enzymatic hydrolysis (cellulase + pectinase) efficiently degrades plant cell wall cellulose and intercellular pectin, significantly improving the cell wall disruption rate. Dual-frequency alternating ultrasound (40kHz / 100kHz), through the synergistic effect of cavitation bubbles at different frequencies, breaks down cell structures of different sizes, promoting the full release of intracellular active components. HPD-100 macroporous resin exhibits excellent selective adsorption capacity for moderately polar phenolic acids and flavonoid glycosides. A gradient elution strategy, using water for impurity removal and 65% ethanol for targeted elution, effectively removes water-soluble impurities such as polysaccharides and proteins, achieving precise capture and enrichment of active components. According to the test results, the total content of the main active ingredients (chlorogenic acid, 1,5-dicaffeoylquinic acid, luteolin-7-O-glucoside) in the extract of this invention is not less than 35%, which is much higher than that of traditional extraction methods.
[0035] Meanwhile, the IC50 of the extract of this invention against electric eel acetylcholinesterase was determined by the Ellman method. 50 The concentration was 18.5 ug / mL, and under the same testing conditions, its activity was equivalent to 65% of the positive control drug donepezil. More importantly, the extract of this invention is derived from natural plants, and its toxicity is far lower than that of chemically synthesized drugs, exhibiting better long-term safety. Animal experiments have shown that the extract of this invention can significantly reduce AChE activity in the brains of mice with scopolamine-induced memory impairment and improve learning and memory function.
[0036] This invention combines extracts with phospholipid materials to form phospholipid complexes. The amphiphilic properties of phospholipids improve the lipid solubility of the extracts, promoting their cross-linking across the blood-brain barrier via passive diffusion or liposome-like mechanisms. Pharmacokinetic experiments show that the phospholipid complex formulation increases the concentration of the extract in mouse brain tissue by 2.3 times compared to the ordinary extract, significantly enhancing drug distribution in the central nervous system and overcoming the bottleneck problem of poor brain penetration of natural extracts. Detailed Implementation
[0037] Example 1: Preparation of Saussurea medusa extract
[0038] Take 1 kg of dried whole plant of Saussurea medusa from Yushu area of Qinghai, crush it and pass it through a 60-mesh sieve to obtain Saussurea medusa coarse powder.
[0039] Step A: Complex enzymatic hydrolysis to break cell walls
[0040] Add 15L of acetate-sodium acetate buffer solution (pH 4.8, material-to-liquid ratio 1:15) to the above coarse powder. Add 12g of compound enzyme (8g cellulase, 4g pectinase, mass ratio 2:1, total enzyme content 1.2%), place the mixture in a 45℃ constant temperature water bath shaker, and shake at 150rpm for 2 hours for enzymatic hydrolysis.
[0041] Step B, Dual-frequency ultrasound-assisted extraction
[0042] The enzymatic hydrolysate was transferred to an ultrasonic extractor, and ultrasonic waves of 40kHz and 100kHz were used alternately, with the frequency switching every 10 minutes. The ultrasonic power was 300W, and the extraction time was 30 minutes.
[0043] Step C: Dynamic adsorption and directional elution of macroporous resin
[0044] The ultrasonic extract was centrifuged at 4000 rpm for 15 minutes, and the supernatant was collected. The supernatant was loaded onto a chromatography column packed with HPD-100 macroporous adsorption resin (resin bed volume approximately 5 L) at a flow rate of 2 BV / h. The column was eluted with 25 L (5 BV) of deionized water at a flow rate of 2.5 BV / h, and the water eluent was discarded. The column was then eluted with 32.5 L (6.5 BV) of 65% ethanol aqueous solution at a flow rate of 1.5 BV / h, and the ethanol eluent was collected.
[0045] Step D, Concentration and Drying
[0046] The ethanol eluent was concentrated under vacuum thin-film treatment at 50℃ and -0.08 MPa until a relative density of 1.10 was obtained (measured at 60℃). The concentrate was then spray-dried with the inlet air temperature controlled at 180℃ and the outlet air temperature at 85℃ to obtain 85.6 g of a pale yellow powder extract, with an extraction rate of 8.56%.
[0047] Extract characterization:
[0048] The content of major active ingredients in the extract was determined by HPLC. Chromatographic conditions: C18 column (250 mm × 4.6 mm, 5 μm); mobile phase: acetonitrile (A) - 0.1% phosphoric acid aqueous solution (B), gradient elution: 0-10 min, 10%-15% A; 10-25 min, 15%-25% A; 25-40 min, 25%-35% A; 40-50 min, 35%-45% A; flow rate: 1.0 mL / min; detection wavelength: 327 nm (chlorogenic acid) and 350 nm (flavonoids); column temperature: 30 ℃; injection volume: 10 μL.
[0049] The extract contained 8.2% chlorogenic acid, 12.5% 1,5-dicaffeoylquinic acid, and 15.8% luteolin-7-O-glucoside, with a total content of 36.5%.
[0050] AChE inhibitory activity assay:
[0051] The inhibitory activity of the extract against acetylcholinesterase in electric eel was determined using the Ellman method. The total reaction volume was 200 μL, containing 0.1 M phosphate buffer (pH 8.0), 0.3 mM DTNB, 0.45 mM ATChI, 0.05 U / mL AChE, and different concentrations of the test sample. After incubation at 37 °C for 15 minutes, the absorbance was measured at 412 nm. Donepezil (10 μM) was used as a positive control.
[0052] The results showed that when the extract concentration was 50 μg / mL, the inhibition rate against AChE was 78.2%; the inhibition rate of donepezil (10 μM), a positive control agent, was 85.5% under the same conditions. The IC50 of the extract against AChE was calculated. 50 The value was 18.5 ug / mL.
[0053] Example 2: Preparation of phospholipid complex
[0054] Take 10g of the extract obtained in Example 1, add 20g of soybean lecithin, dissolve in 200mL of anhydrous ethanol, and stir at 200rpm for 1 hour in a 40℃ water bath. Place the resulting solution in a rotary evaporator and remove the ethanol by rotary evaporation under reduced pressure at 40℃ to obtain 28.5g of a pale yellow solid dispersion, namely the phospholipid complex of the jellyfish snow lotus extract.
[0055] The above phospholipid complex was mixed with excipients such as microcrystalline cellulose, lactose, and magnesium stearate according to conventional tablet manufacturing processes and compressed into tablets containing 100 mg of extract per tablet.
[0056] Example 3: Pharmacodynamic Experiment of Scopolamine-Induced Memory Impairment Model in Mice
[0057] Laboratory animals: SPF-grade male ICR mice, weighing 18-22g, provided by the laboratory animal center, acclimatized for 7 days, with free access to water and food.
[0058] Experimental grouping and drug administration: Mice were randomly divided into 5 groups, with 10 mice in each group:
[0059] Sham surgery group: Administered an equal volume of normal saline via gavage for 14 consecutive days;
[0060] Model group: Administered an equal volume of physiological saline by gavage for 14 consecutive days;
[0061] Positive drug group: donepezil 5 mg / kg was administered by gavage for 14 consecutive days;
[0062] Extract group: The extract obtained in Example 1 was administered by gavage at a dose of 100 mg / kg for 14 consecutive days;
[0063] Phospholipid complex group: The phospholipid complex obtained in Example 2 (equivalent to 100 mg / kg of extract) was administered by gavage for 14 consecutive days.
[0064] Except for the sham surgery group, the other groups were given intraperitoneal injections of scopolamine 1 mg / kg on days 8-14 of drug administration to establish a memory impairment model, while the sham surgery group was given an intraperitoneal injection of an equal volume of physiological saline.
[0065] Morris water maze test: The Morris water maze test was conducted 1 hour after the last drug administration. The water maze pool was 120 cm in diameter and 30 cm deep, with a water temperature of 25 ± 1℃. The platform was 10 cm in diameter, placed in the center of the third quadrant, and submerged 1 cm below the water surface. The orientation and navigation experiment lasted for 5 days, with training 4 times a day. The escape latency of the mice when they found the platform was recorded. On the 6th day, the platform was removed, and a spatial exploration experiment was conducted. The number of times the mice crossed the original platform location within 60 seconds was recorded.
[0066] Assay for AChE activity in brain tissue: After the behavioral experiment, mice were decapitated and their brains were removed. The cerebral cortex and hippocampus were separated and mixed with pre-cooled physiological saline to prepare a 10% tissue homogenate. The AChE activity in the brain tissue was measured according to the instructions of the acetylcholinesterase kit. The protein concentration was determined using the BCA method.
[0067] Experimental results:
[0068] Group Dosage (mg / kg) Avoiding the incubation period (s) Platform crossing times Brain AChE activity (u / mg prot) Sham surgery group - 15.2±3.1 4.8±0.9 0.85±0.07 Model group - 58.6±5.2 0.5±0.2 2.15±0.12 Positive drug group 5 22.4±4.1 3.5±0.8 1.25±0.09 Extract group 100 28.7±4.5 2.9±0.7 1.42±0.11 Phospholipid complex group 100 19.8±3.5 4.2±0.8 1.05±0.08
[0069] Note: Compared with the sham surgery group, ##p<0.01; compared with the model group, *p<0.05, **p<0.01; compared with the extract group, △p<0.05.
[0070] Results analysis:
[0071] Compared with the sham-operated group, the escape latency of mice in the model group was significantly prolonged (p<0.01), the platform crossing coefficient was significantly reduced (p<0.01), and the AChE activity in the brain was significantly increased (p<0.01), indicating that the scopolamine-induced memory impairment model was successfully established.
[0072] Compared with the model group, the positive control group, donepezil group, extract group and phospholipid complex group all significantly shortened the escape latency, increased the number of platform crossings and reduced AChE activity in the brain (all p<0.01), indicating that the extract of the present invention has the effect of improving learning and memory impairment and inhibiting AChE activity in the brain.
[0073] Of particular note is that the escape latency in the phospholipid complex group (19.8±3.5s) was significantly shorter than that in the ordinary extract group (28.7±4.5s, p<0.05), the number of platform crossings (4.2±0.8 times) was significantly greater than that in the ordinary extract group (2.9±0.7 times, p<0.05), and the intracranial AChE activity (1.05±0.08 U / mg prot) was significantly lower than that in the ordinary extract group (1.42±0.11 U / mg prot, p<0.05), with no significant difference compared to the positive control group (donepezil) and the sham operation group. These results fully demonstrate that the phospholipid complex technology significantly enhances the extract's ability to cross the blood-brain barrier, enabling it to exert a stronger AChE inhibitory effect and cognitive-improving effect in the central nervous system.
[0074] Example 4: Comparative Experiment of Different Extraction Methods
[0075] To verify the technical advantages of the integrated process of this invention, the following comparative experiments were conducted:
[0076] Comparative Example 1 (Traditional Water Decoction Method): Take 1 kg of dried snow lotus, add 10 times the amount of water, decoct twice, 1 hour each time, combine the decoctions, concentrate and dry.
[0077] Comparative Example 2 (Ordinary alcohol extraction method): Take 1 kg of dried snow lotus, add 10 times the amount of 70% ethanol, reflux extract twice, 1.5 hours each time, combine the extractors, recover the ethanol, concentrate and dry.
[0078] Comparative Example 3 (Enzyme-free single-frequency ultrasonic extraction): The steps of Example 1 were followed, but the enzymatic hydrolysis process in step A was omitted. Single-frequency ultrasonic extraction at 100 kHz was used for 30 minutes. Subsequent steps were the same as in Example 1.
[0079] Comparative Example 4 (without macroporous resin purification): Followed steps A and B of Example 1, but omitted step C, macroporous resin purification, and directly concentrated and dried the ultrasonic extract.
[0080] The content of active ingredients and AChE inhibitory activity of the extracts obtained from each group were determined according to the method in Example 1. The results are as follows:
[0081] Extraction method Extraction yield Total content of three types of active ingredients AChE inhibition rate (50ug / mL) <![CDATA[IC 50 (μg / mL)]]> Example 1 8.56% 36.5% 78.2% 18.5 Comparative Example 1 18.2% 8.3% 32.5% 125.6 Comparative Example 2 14.5% 12.6% 41.8% 86.4 Comparative Example 3 11.3% 22.4% 55.3% 52.7 Comparative Example 4 10.8% 15.2% 48.6% 68.3
[0082] The results show that the integrated process of Example 1 of the present invention is significantly better than the comparative examples in terms of active ingredient enrichment and AChE inhibitory activity, proving that the three-step synergistic process of compound enzymatic hydrolysis, dual-frequency ultrasound and macroporous resin purification has significant technological progress.
[0083] Although embodiments of the invention have been shown and described, it will be understood by those skilled in the art that various changes, modifications, substitutions and alterations can be made to these embodiments without departing from the principles and spirit of the invention, the scope of which is defined by the appended embodiments and their equivalents.
Claims
1. A method for preparing a jellyfish snow lotus extract with acetylcholinesterase inhibitory activity, characterized in that, Includes the following steps: Step A: Complex enzymatic hydrolysis to break cell walls Take the dried whole plant of Saussurea medusa, crush and sieve it to obtain Saussurea medusa coarse powder; add an acetate-sodium acetate buffer solution with a pH of 4.5-5.0 to the coarse powder, the amount of the buffer solution added is based on a material-to-liquid ratio of 1:10-1:20 (w / v); add a compound enzyme to the above mixture, the compound enzyme is composed of cellulase and pectinase, the mass ratio of the two is 1.5-2.5:1, and the total amount of the compound enzyme added is 0.8%-1.5% of the mass of the coarse powder; place the above mixture in a constant temperature water bath at 40℃-50℃, shake and enzymatically hydrolyze for 1.5-2.5 hours to obtain the enzymatic hydrolysate; Step B, Dual-frequency ultrasound-assisted extraction The enzymatic hydrolysate obtained in step A was placed in an ultrasonic extraction device and extracted using a dual-frequency alternating ultrasonic method of 40kHz and 100kHz, switching the frequency every 8-12 minutes. The ultrasonic power was 200W-400W, and the extraction time was 25-35 minutes to obtain the extract. Step C: Dynamic adsorption and directional elution of macroporous resin The extract obtained in step B is subjected to solid-liquid separation, and the supernatant is collected. The supernatant is loaded onto a chromatography column packed with HPD-100 macroporous adsorption resin at a flow rate of 1.5-2.5 BV / h for dynamic adsorption. After adsorption, 4-6 BV of deionized water is used to elute at a flow rate of 2-3 BV / h to remove water-soluble impurities. Then, 60%-70% ethanol aqueous solution is used as the eluent and eluted at a flow rate of 1.0-2.0 BV / h. The ethanol eluent is collected. Step D, Concentration and Drying The ethanol eluent collected in step C was concentrated under vacuum thin-film conditions at 45℃-55℃ and a vacuum degree of -0.06-0.09 MPa until the relative density was 1.05-1.15, resulting in a concentrate. The concentrate was then spray-dried with the inlet air temperature controlled at 170℃-190℃ and the outlet air temperature controlled at 80℃-90℃ to obtain the jellyfish snow lotus extract.
2. The method for preparing a *Saussurea involucrata* extract with acetylcholinesterase inhibitory activity according to claim 1, characterized in that: In step A, the pulverization and sieving are performed through a 40-80 mesh sieve, the pH value is 4.8, the material-to-liquid ratio is 1:15, the mass ratio of cellulase to pectinase is 2:1, the total enzyme content is 1.2%, the enzymatic hydrolysis temperature is 45℃, and the enzymatic hydrolysis time is 2 hours.
3. The method for preparing a *Saussurea involucrata* extract with acetylcholinesterase inhibitory activity according to claim 1, characterized in that: In step B, the dual-frequency alternating ultrasound switches frequencies every 10 minutes, the ultrasound power is 300W, and the extraction time is 30 minutes.
4. The method for preparing a *Saussurea involucrata* extract with acetylcholinesterase inhibitory activity according to claim 1, characterized in that: In step C, the sample loading flow rate is 2 BV / h, the deionized water elution volume is 5 BV and the flow rate is 2.5 BV / h, and the volume fraction of the ethanol aqueous solution is 65% with an elution flow rate of 1.5 BV / h.
5. The method for preparing a *Saussurea involucrata* extract with acetylcholinesterase inhibitory activity according to claim 1, characterized in that: In step D, the concentration temperature is 50°C, the vacuum degree is -0.08 MPa, the relative density is 1.10, the inlet air temperature for spray drying is 180°C, and the outlet air temperature is 85°C.
6. A jellyfish extract with acetylcholinesterase inhibitory activity, characterized in that, The extract is prepared by any one of claims 1 to 5, and contains chlorogenic acid, 1,5-dicaffeoylquinic acid and luteolin-7-O-glucoside, with the total mass fraction of the three active ingredients being not less than 35%.
7. The jellyfish extract with acetylcholinesterase inhibitory activity according to claim 6, characterized in that, The extract contains at least 15% chlorogenic acid and at least 10% flavonoid glycosides by mass.
8. The jellyfish extract with acetylcholinesterase inhibitory activity according to claim 6, characterized in that, The extract has a half-maximal inhibitory concentration (IC50) against electric eel acetylcholinesterase. 50 The value is 15-22 ug / mL.
9. A pharmaceutical composition, characterized in that, The extract includes the jellyfish snow lotus extract according to any one of claims 6 to 8 and a pharmaceutically acceptable carrier, the carrier comprising a gelling material, wherein the extract and the gelling material form a gelling complex.
10. The pharmaceutical composition according to claim 9, characterized in that, The extract and soybean egg curd were dissolved in anhydrous ethanol at a mass ratio of 1:1.5-2.5, and stirred at 35℃-45℃. The organic solvent was then removed by rotary evaporation to form a solid dispersion, namely the curd complex.
11. The pharmaceutical composition according to claim 10, characterized in that, The mass ratio of the extract to soybean ovoid is 1:2, and the dissolution temperature is 40°C.
12. The pharmaceutical composition according to claim 9, characterized in that, The dosage form of the pharmaceutical composition is capsules, tablets, granules or pills.
13. The jellyfish extract according to any one of claims 6 to 8 or the pharmaceutical composition according to any one of claims 9 to 12, characterized in that, The application described in the preparation of medicaments for the prevention and / or treatment of Alzheimer's disease.
14. A pharmaceutical composition according to claim 9, characterized in that, The drug exerts its effects on Alzheimer's disease by inhibiting alcohol cholinesterase activity, increasing acetylcholine levels in the brain, and improving learning and memory impairments.