CN114848689B - Herba Lycopodii Serrati effective component, its preparation method and application in preparing medicine for preventing or treating senile dementia - Google Patents
Herba Lycopodii Serrati effective component, its preparation method and application in preparing medicine for preventing or treating senile dementia Download PDFInfo
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Abstract
The invention discloses a huperzia serrata effective part, a preparation method thereof and application thereof in preparing a medicament with the effect of preventing or treating senile dementia. The preparation method comprises the following steps: (1) Extracting herba Lycopodii Serrati with ethanol, concentrating the extractive solution, and removing ethanol to obtain herba Lycopodii Serrati total extract; (2) Adding water into the oroxylum indicum total extract for suspension, then adding acid to adjust the pH value of the whole suspension system to 2-3, standing, and then adding an organic solvent A for extraction to obtain an organic solvent A layer and an acid water layer; (3) Taking an acid water layer, adding alkali into the acid water layer, adjusting the pH value of the acid water layer to 9-10, standing, and adding an organic solvent B for extraction to obtain an organic solvent B layer and an alkali water layer; concentrating and drying the organic solvent layer B to obtain the effective part of the huperzia serrata. The effective part of the huperzia serrata prepared by the method has excellent acetylcholinesterase inhibition effect; therefore, the application of the compound has wide application prospect in developing medicaments for preventing and treating senile dementia.
Description
Technical Field
The invention relates to the technical field of natural medicinal chemistry, in particular to a huperzia serrata effective part, a preparation method thereof and application thereof in preparing a medicament with an effect of preventing or treating senile dementia.
Background
Herba Lycopodii Serrati is herba Huperzia serrata of huperziaceae of huperzia, and is prepared from whole herb; it has effects in dispelling blood stasis, stopping bleeding, clearing away heat and toxic materials, and relieving swelling and pain; can be used for treating traumatic injury, swelling and pain due to blood stasis, internal injury, and hematemesis; it is used externally to treat carbuncle, furuncle, toxic swelling, snake bite, burn and scald.
Alzheimer's Disease (AD) is a progressive degenerative disease of the nervous system with occult disease; patients who are older than 65 years are called presenile dementia; the patient who is aged 65 is called senile dementia. The acetylcholinesterase inhibitor, also called anticholinesterase drug, achieves the accumulation of acetylcholine (ACh) at synapse, prolongs and increases the action of acetylcholine by reversible inhibition of acetylcholinesterase; is widely used for resisting senile dementia clinically.
Therefore, the development of a drug having an inhibitory effect on acetylcholinesterase is one of the directions for the development of a drug having an effect of preventing or treating senile dementia. In the prior art, a monomer compound huperzine A is separated from huperzia serrata, and researches show that the huperzine A has an obvious inhibition effect on acetylcholinesterase and has a remarkable curative effect on senile dementia.
However, there are few reports in the prior art on whether other effective components in huperzia serrata have acetylcholinesterase inhibitory effect. Therefore, the effective part or monomer compound with the inhibitory effect on acetylcholinesterase is further developed by taking the traditional Chinese medicine huperzia serrata as the raw material, more choices are provided for developing the medicine for preventing or treating the senile dementia, and the huperzia serrata has important application value.
Disclosure of Invention
In order to overcome at least one technical problem in the prior art, the invention firstly provides a preparation method of the effective part of the huperzia serrata. Research shows that the effective part of the huperzia serrata prepared by the method has excellent acetylcholinesterase inhibition effect.
The invention aims to solve the technical problems and is realized by the following technical scheme:
a preparation method of effective parts of huperzia serrata is characterized by comprising the following steps:
(1) Extracting herba Lycopodii Serrati with ethanol, concentrating the extractive solution, and removing ethanol to obtain herba Lycopodii Serrati total extract;
(2) Adding water into the oroxylum indicum total extract for suspension, then adding acid to adjust the pH value of the whole suspension system to 2-3, standing, and then adding an organic solvent A for extraction to obtain an organic solvent A layer and an acid water layer;
(3) Taking an acid water layer, adding alkali into the acid water layer, adjusting the pH value of the acid water layer to 9-10, standing, and adding an organic solvent B for extraction to obtain an organic solvent B layer and an alkali water layer; concentrating and drying the organic solvent layer B to obtain the effective part of the huperzia serrata.
Preferably, the ethanol in the step (1) refers to 70-95% by volume of ethanol aqueous solution.
Preferably, the dosage ratio of the huperzia serrata to the ethanol in the step (1) is 1g.
Most preferably, the usage ratio of the huperzine serrate to the ethanol in the step (1) is 1g.
Preferably, the extraction in step (1) is cold soaking extraction, and the extraction time is 3-10 days.
Most preferably, the extraction time is 7d.
Preferably, the dosage ratio of the total extractum of the oroxylum indicum to water in the step (2) is 1g.
Most preferably, the dosage ratio of the total extract of the oroxylum indicum to the water in the step (2) is 1g.
Preferably, the organic solvent A in the step (2) is dichloromethane; in the step (3), the organic solvent B is dichloromethane.
The invention provides a brand-new preparation method of the effective part of the huperzia serrata; the inventors found in a large number of experiments that the pH control of steps (2) and (3) of the method and the selection of organic solvent a and organic solvent B are an integrated solution, which together play a decisive role in determining whether the prepared effective part of huperzia serrata has excellent acetylcholinesterase inhibition.
The inventors have found in a great deal of research that: when the pH value in the step (2) is controlled to be 2-3, the pH value in the step (3) is controlled to be 9-10, and the organic solvents A and B simultaneously select dichloromethane, the prepared effective part of the huperzia serrata has excellent acetylcholinesterase inhibition effect. If the pH values of the steps (2) and (3) are in other ranges, or the organic solvents A and B are other organic solvents, the inhibitory effect of the effective parts of the huperzia serrata prepared by the method on the acetylcholinesterase is obviously smaller than that of the effective parts of the huperzia serrata prepared by the method, and even the effective parts with the acetylcholinesterase inhibitory effect cannot be prepared.
Preferably, the organic solvent A in the step (2) is dichloromethane; the organic solvent B in the step (3) is a mixed solvent consisting of dichloromethane and diethyl ether.
The inventor surprisingly found in further research that: when the pH value in the step (2) is controlled to be 2-3, the pH value in the step (3) is controlled to be 9-10, the organic solvent A is dichloromethane, and the organic solvent B is a combination of dichloromethane and diethyl ether, the prepared effective part of the huperzia serrata has more excellent acetylcholinesterase inhibition effect; compared with the effective part of the huperzia serrata prepared by only selecting dichloromethane as the organic solvent B, the inhibition effect of the huperzia serrata on acetylcholinesterase is further greatly improved.
Wherein the volume ratio of the dichloromethane to the ether is 4-6; most preferably, the volume ratio of dichloromethane to diethyl ether is 5.
Preferably, the volume dosage of the organic solvent A in the step (2) and the volume dosage of the water are in a ratio of 1-2; the volume ratio of the organic solvent B to the acid water layer in the step (3) is 1-2.
Preferably, the acid in step (2) is concentrated hydrochloric acid; the alkali in the step (3) is ammonia water.
The invention also provides the effective part of the huperzia serrata prepared by the preparation method.
The invention also provides application of the effective part of the huperzia serrata in preparing an acetylcholinesterase inhibitor.
The invention also provides application of the effective part of the huperzia serrata in preparing a medicament with the effect of preventing or treating senile dementia.
Has the advantages that: the invention provides a brand-new preparation method of the effective part of the huperzia serrata, and the effective part of the huperzia serrata prepared by the method has excellent acetylcholinesterase inhibition effect. Therefore, the effective part of the huperzia serrata has wide application prospect in developing the medicine for preventing and treating the senile dementia.
Detailed Description
The present invention will be further explained with reference to specific examples, which are not intended to limit the present invention in any way.
Example 1 preparation of effective fractions of Huperzia serrata
(1) Extracting herba Lycopodii Serrati with ethanol (95 vol% ethanol water solution) at room temperature for 7 days, concentrating the extractive solution, and removing ethanol to obtain herba Lycopodii Serrati total extract;
wherein the dosage ratio of the huperzia serrata to the ethanol is 1g;
(2) Adding water into the oroxylum indicum total extract for suspension, then adding concentrated hydrochloric acid to adjust the pH value of the whole suspension system to 2.5, standing for 10 hours, and then adding an organic solvent A for extraction to obtain an organic solvent A layer and an acid water layer;
wherein the dosage ratio of the oroxylum indicum total extract to water is 1g to 20mL; the volume dosage ratio of the organic solvent A to the water is 2; the organic solvent A is dichloromethane;
(3) Taking an acid water layer, adding ammonia water into the acid water layer, adjusting the pH value of the acid water layer to 9.5, standing, and adding an organic solvent B for extraction to obtain an organic solvent B layer and an alkali water layer; concentrating and drying the organic solvent layer B to obtain the effective part of the huperzia serrata;
wherein the volume ratio of the organic solvent B to the acid water layer is 2; the organic solvent B is dichloromethane.
Example 2 preparation of effective fractions of Huperzia serrata
(1) Extracting herba Lycopodii Serrati with ethanol (70 vol% ethanol water solution) at room temperature for 10 days, concentrating the extractive solution, and removing ethanol to obtain herba Lycopodii Serrati total extract;
wherein the using amount ratio of the huperzia serrata to the ethanol is 1g;
(2) Adding water into the oroxylum indicum total extract for suspension, then adding concentrated hydrochloric acid to adjust the pH value of the whole suspension system to 2, standing for 10 hours, and then adding an organic solvent A for extraction to obtain an organic solvent A layer and an acid water layer;
wherein the dosage ratio of the total extract of the oroxylum indicum to the water is 1g; the volume dosage of the organic solvent A and the volume dosage of the water are in a ratio of 2; the organic solvent A is dichloromethane;
(3) Taking an acid water layer, adding ammonia water into the acid water layer, adjusting the pH value of the acid water layer to 9, standing, and adding an organic solvent B for extraction to obtain an organic solvent B layer and an alkali water layer; concentrating and drying the organic solvent layer B to obtain the effective part of the huperzia serrata;
wherein the volume ratio of the organic solvent B to the acid water layer is 2; the organic solvent B is dichloromethane.
Example 3 preparation of effective fractions of Huperzia serrata
(1) Extracting herba Lycopodii Serrati with ethanol (80 vol% ethanol water solution) at room temperature for 5 days, concentrating the extractive solution, and removing ethanol to obtain herba Lycopodii Serrati total extract;
wherein the dosage ratio of the huperzia serrata to the ethanol is 1g;
(2) Adding water into the oroxylum indicum total extract for suspension, then adding concentrated hydrochloric acid to adjust the pH value of the whole suspension system to 3, standing for 10 hours, and then adding an organic solvent A for extraction to obtain an organic solvent A layer and an acid water layer;
wherein the dosage ratio of the total extract of the oroxylum indicum to the water is 1g; the volume dosage of the organic solvent A and the volume dosage of the water are in a ratio of 1; the organic solvent A is dichloromethane;
(3) Taking an acid water layer, adding ammonia water into the acid water layer, adjusting the pH value of the acid water layer to 10, standing, and adding an organic solvent B for extraction to obtain an organic solvent B layer and an alkali water layer; concentrating and drying the organic solvent layer B to obtain the effective part of the huperzia serrata;
wherein the volume ratio of the organic solvent B to the acid water layer is 1; the organic solvent B is dichloromethane.
Example 4 preparation of effective fractions of Huperzia serrata
(1) Extracting herba Lycopodii Serrati with ethanol (95 vol% ethanol water solution) at room temperature for 7 days, concentrating the extractive solution, and removing ethanol to obtain herba Lycopodii Serrati total extract;
wherein the dosage ratio of the huperzia serrata to the ethanol is 1g;
(2) Adding water into the oroxylum indicum total extract for suspension, then adding concentrated hydrochloric acid to adjust the pH value of the whole suspension system to 2.5, standing for 10 hours, and then adding an organic solvent A for extraction to obtain an organic solvent A layer and an acid water layer;
wherein the dosage ratio of the total extract of the huperzia serrata to water is 1g; the volume dosage of the organic solvent A and the volume dosage of the water are in a ratio of 2; the organic solvent A is dichloromethane;
(3) Taking an acid water layer, adding ammonia water into the acid water layer, adjusting the pH value of the acid water layer to 9.5, standing, and adding an organic solvent B for extraction to obtain an organic solvent B layer and an alkaline water layer; concentrating and drying the organic solvent layer B to obtain the effective part of the huperzia serrata;
wherein the volume ratio of the organic solvent B to the acid water layer is 2; the organic solvent B is a mixed organic solvent consisting of dichloromethane and diethyl ether with a volume ratio of 5.
Comparative example 1 preparation of effective fraction of Huperzia serrata
(1) Extracting herba Lycopodii Serrati with ethanol (95 vol% ethanol water solution) at room temperature for 7 days, concentrating the extractive solution, and removing ethanol to obtain herba Lycopodii Serrati total extract;
wherein the dosage ratio of the huperzia serrata to the ethanol is 1g;
(2) Adding water into the oroxylum indicum total extract for suspension, then adding concentrated hydrochloric acid to adjust the pH value of the whole suspension system to 4, standing for 10 hours, and adding an organic solvent A for extraction to obtain an organic solvent A layer and an acid water layer;
wherein the dosage ratio of the total extract of the huperzia serrata to water is 1g; the volume dosage of the organic solvent A and the volume dosage of the water are in a ratio of 2; the organic solvent A is chloroform;
(3) Taking an acid water layer, adding ammonia water into the acid water layer, adjusting the pH value of the acid water layer to 11, standing, and adding an organic solvent B for extraction to obtain an organic solvent B layer and an alkali water layer; concentrating and drying the organic solvent layer B to obtain the effective part of the huperzia serrata;
wherein the volume ratio of the organic solvent B to the acid water layer is 2; and the organic solvent B is chloroform.
Comparative example 1 differs from example 1 in that the pH values in steps (2) and (3) are different, and in that the organic solvents a and B are different; comparative example 1 the pH in step (2) was controlled to 4, the pH in step (3) was controlled to 11, and chloroform was used as both organic solvents a and B; in example 1, the pH value in step (2) is controlled to be 2.5, the pH value in step (3) is controlled to be 9.5, and dichloromethane is simultaneously used as the organic solvents A and B.
Comparative example 2 preparation of effective fraction of Huperzia serrata
(1) Extracting herba Lycopodii Serrati with ethanol (95 vol% ethanol water solution) at room temperature for 7 days, concentrating the extractive solution, and removing ethanol to obtain herba Lycopodii Serrati total extract;
wherein the dosage ratio of the huperzia serrata to the ethanol is 1g;
(2) Adding water into the oroxylum indicum total extract for suspension, then adding concentrated hydrochloric acid to adjust the pH value of the whole suspension system to 4, standing for 10 hours, and adding an organic solvent A for extraction to obtain an organic solvent A layer and an acid water layer;
wherein the dosage ratio of the total extract of the huperzia serrata to water is 1g; the volume dosage of the organic solvent A and the volume dosage of the water are in a ratio of 2; the organic solvent A is ethyl acetate;
(3) Taking an acid water layer, adding ammonia water into the acid water layer, adjusting the pH value of the acid water layer to 11, standing, and adding an organic solvent B for extraction to obtain an organic solvent B layer and an alkali water layer; concentrating and drying the organic solvent layer B to obtain the effective part of the huperzia serrata;
wherein the volume ratio of the organic solvent B to the acid water layer is 2; the organic solvent B is ethyl acetate.
Comparative example 1 differs from example 1 in that the pH values in steps (2) and (3) are different, and in that the organic solvents a and B are different; comparative example 1 the pH in step (2) was controlled at 4, the pH in step (3) was controlled at 11, and both organic solvents a and B were ethyl acetate; in example 1, the pH value in step (2) was controlled to 2.5, the pH value in step (3) was controlled to 9.5, and ethyl acetate was used as the organic solvents A and B.
Comparative example 3 preparation of effective fraction of Huperzia serrata
(1) Extracting herba Lycopodii Serrati with ethanol (95 vol% ethanol water solution) at room temperature for 7 days, concentrating the extractive solution, and removing ethanol to obtain herba Lycopodii Serrati total extract;
wherein the dosage ratio of the huperzia serrata to the ethanol is 1g;
(2) Adding water into the oroxylum indicum total extract for suspension, then adding concentrated hydrochloric acid to adjust the pH value of the whole suspension system to 2.5, standing for 10 hours, and then adding an organic solvent A for extraction to obtain an organic solvent A layer and an acid water layer;
wherein the dosage ratio of the oroxylum indicum total extract to water is 1g to 20mL; the volume dosage ratio of the organic solvent A to the water is 2; the organic solvent A is dichloromethane;
(3) Taking an acid water layer, adding ammonia water into the acid water layer, adjusting the pH value of the acid water layer to 9.5, standing, and adding an organic solvent B for extraction to obtain an organic solvent B layer and an alkaline water layer; concentrating and drying the organic solvent layer B to obtain the effective part of the huperzia serrata;
wherein the volume ratio of the organic solvent B to the acid water layer is 2; the organic solvent B is a mixed organic solvent consisting of dichloromethane and ethyl acetate with a volume ratio of 5.
Comparative example 3 is different from example 4 in that the organic solvent B is selected differently, and the organic solvent B in comparative example 3 is a mixed organic solvent consisting of dichloromethane and ethyl acetate; and the organic solvent B in the embodiment 4 is a mixed organic solvent consisting of dichloromethane and diethyl ether.
Comparative example 4 preparation of effective fraction of Huperzia serrata
(1) Extracting herba Lycopodii Serrati with ethanol (95 vol% ethanol water solution) at room temperature for 7 days, concentrating the extractive solution, and removing ethanol to obtain herba Lycopodii Serrati total extract;
wherein the dosage ratio of the huperzia serrata to the ethanol is 1g;
(2) Adding water into the oroxylum indicum total extract for suspension, then adding concentrated hydrochloric acid to adjust the pH value of the whole suspension system to 2.5, standing for 10 hours, and then adding an organic solvent A for extraction to obtain an organic solvent A layer and an acid water layer;
wherein the dosage ratio of the oroxylum indicum total extract to water is 1g to 20mL; the volume dosage of the organic solvent A and the volume dosage of the water are in a ratio of 2; the organic solvent A is dichloromethane;
(3) Taking an acid water layer, adding ammonia water into the acid water layer, adjusting the pH value of the acid water layer to 9.5, standing, and adding an organic solvent B for extraction to obtain an organic solvent B layer and an alkali water layer; concentrating and drying the organic solvent layer B to obtain the effective part of the huperzia serrata;
wherein the volume ratio of the organic solvent B to the acid water layer is 2; the organic solvent B is a mixed organic solvent consisting of dichloromethane and cyclohexane in a volume ratio of 5.
Comparative example 4 is different from example 4 in that the organic solvent B is selected differently, and the organic solvent B in comparative example 4 is a mixed organic solvent composed of dichloromethane and cyclohexane; and the organic solvent B in the embodiment 4 is a mixed organic solvent consisting of dichloromethane and diethyl ether.
Examples of the experiments
Dissolving the effective parts of the huperzia serrata prepared in the embodiments 1-4 and the comparative examples 1-4 by using DMSO (dimethyl sulfoxide), and preparing a solution to be detected, wherein the solution to be detected is 2 mg/mL; the method is used for testing the inhibition rate of the effective part of the huperzia serrata prepared in examples 1-4 and comparative examples 1-4 on the acetylcholine esterase.
Taking a 96-well plate, and respectively adding 10 mu L of solution to be detected, 40 mu L of Tris-HCl buffer solution with the pH value of 8.0 and 10 mu L of acetylcholinesterase solution (prepared by using Tris-HCl buffer solution with the pH value of 8.0, and the concentration of the Tris-HCl buffer solution is 1000U/mL) into each well; the blank control group replaces the solution to be detected with an equivalent amount of Tris-HCl buffer solution with the pH value of 8.0; then incubating in a shaker at 37 ℃ for 10min; then 20. Mu.L of color development reagent DTNB solution (prepared by Tris-HCl buffer solution with pH value of 8.0, concentration of 0.1 mM) and 10. Mu.L of substrate acetylthiocholine iodide solution (prepared by Tris-HCl buffer solution with pH value of 8.0, concentration of 0.2 mM) are added, and then the incubation is continued in a shaking table for 10min at 37 ℃; after the incubation is finished, measuring the absorbance under 412nm by using an enzyme-labeling instrument; calculating the inhibition rate of the solution to be detected on acetylcholinesterase; the test results are shown in Table 1.
TABLE 1 inhibitory effect of effective fractions of Huperzia serrata of the present invention on acetylcholinesterase
As can be seen from the experimental data in Table 1, the effective fraction of huperzia serrata prepared in examples 1-3 has an acetylcholinesterase inhibition rate of more than 60% at a concentration of 2 mg/mL. This indicates that: the effective part of the huperzia serrata prepared by the method has excellent acetylcholinesterase inhibition effect.
As can be seen from the experimental data in Table 1, the effective fraction of huperzia serrata prepared in example 1 has a much higher inhibitory rate to acetylcholinesterase than the effective fraction of huperzia serrata prepared in comparative examples 1 and 2. In contrast, comparative examples 1 and 2 differ from example 1 in the difference in the pH values of steps (2) and (3) and in the difference in the organic solvents A and B; this indicates that: the pH value control of the steps (2) and (3) of the method and the selection of the organic solvent A and the organic solvent B are an integral scheme, and the scheme plays a decisive role in determining whether the prepared effective part of the huperzia serrata has excellent acetylcholinesterase inhibition effect or not; the above studies show that: only when the pH value in the step (2) is controlled to be 2-3, the pH value in the step (3) is controlled to be 9-10, and the organic solvents A and B simultaneously select dichloromethane, the prepared effective part of the huperzia serrata has excellent acetylcholinesterase inhibition effect; the effective parts of the huperzia serrata prepared in the steps (2) and (3) under other pH values and in the organic solvents A and B adopting other organic solvents do not have excellent acetylcholinesterase inhibition effect.
As can be seen from the experimental data in Table 1, the effective part of huperzia serrata prepared in example 4 has an acetylcholinesterase inhibition rate of 91.8% at a concentration of 2 mg/mL; the effective part of huperzia serrata prepared in example 1 is further greatly improved, and the huperzia serrata has excellent acetylcholinesterase inhibition effect. This indicates that: when the pH value in the step (2) is controlled to be 2-3, the pH value in the step (3) is controlled to be 9-10, the organic solvent A is dichloromethane, and the organic solvent B is a combination of dichloromethane and diethyl ether, the prepared effective part of the huperzia serrata has more excellent acetylcholinesterase inhibition effect; compared with the effective part of the huperzia serrata prepared by only selecting dichloromethane as the organic solvent B, the acetylcholinesterase inhibition effect of the huperzia serrata is further and greatly improved.
As can be seen from the experimental data in Table 1, the effective parts of huperzia serrata prepared in comparative examples 3 and 4 have no obvious improvement or even a reduction in the acetylcholinesterase inhibition effect compared with example 1; this indicates that: the inhibition effect of the prepared effective part of the huperzia serrata on acetylcholinesterase can be further greatly improved only when the organic solvent B is selected from a combination of dichloromethane and ether, so that the prepared effective part of the huperzia serrata has more excellent acetylcholinesterase inhibition effect; the organic solvent B is selected from other organic solvents, so that the inhibition effect of the prepared effective part of the huperzia serrata on acetylcholinesterase cannot be improved or remarkably improved.
Claims (8)
1. A preparation method of effective parts of huperzia serrata is characterized by comprising the following steps:
(1) Extracting herba Lycopodii Serrati with ethanol, concentrating the extractive solution, and removing ethanol to obtain herba Lycopodii Serrati total extract;
(2) Adding water into the oroxylum indicum total extract for suspension, then adding acid to adjust the pH value of the whole suspension system to 2-3, standing, and adding an organic solvent A for extraction to obtain an organic solvent A layer and an acid water layer;
(3) Taking an acid water layer, adding alkali into the acid water layer, adjusting the pH value of the acid water layer to 9-10, standing, and adding an organic solvent B for extraction to obtain an organic solvent B layer and an alkali water layer; concentrating and drying the organic solvent layer B to obtain the effective part of the huperzia serrata;
the ethanol in the step (1) is ethanol water solution with volume fraction of 70 to 95 percent;
the dosage ratio of the huperzia serrata to the ethanol in the step (1) is 1g to 20mL;
the extraction in the step (1) is carried out by adopting a cold soaking method, and the extraction time is 3-10d;
the dosage ratio of the total extract of the oroxylum indicum to the water in the step (2) is 1g;
in the step (2), the organic solvent A is dichloromethane;
the volume usage of the organic solvent A and the volume usage of the water in the step (2) are in a ratio of 1 to 2;
in the step (3), the organic solvent B is a mixed solvent consisting of dichloromethane and diethyl ether; wherein the volume ratio of dichloromethane to ether is 4 to 6;
in the step (3), the volume ratio of the organic solvent B to the acid water layer is 1 to 2.
2. The method according to claim 1, wherein the ratio of the amount of huperzine serrate to ethanol used in step (1) is 1g.
3. The method of claim 1, wherein the extraction time is 7 days.
4. The preparation method according to claim 1, wherein the using amount ratio of the oroxylum indicum total extract to the water in the step (2) is 1g to 20mL.
5. The process according to claim 1, wherein the volume ratio of dichloromethane to diethyl ether in the organic solvent B is 5.
6. The effective part of huperzia serrata prepared by the preparation method according to any one of claims 1 to 5.
7. Use of the effective fraction of huperzia serrata of claim 6 for preparing acetylcholinesterase inhibitor.
8. The use of the effective fraction of huperzia serrata of claim 6 for preparing a medicament for preventing or treating senile dementia.
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