一种ERCP专用胆道痉挛缓解复合物、制剂及其制法

本发明提供了一种ERCP专用胆道痉挛缓解复合物、制剂及其制法，该复合物包括质量比为1‑3:13‑25的L‑薄荷醇、β‑环糊精或其衍生物，制剂包括L‑薄荷醇水溶液和L‑薄荷醇纳米乳剂，其中L‑薄荷醇水溶液包括复合物和水，L‑薄荷醇纳米乳包括L‑薄荷醇、表面活性剂和消泡剂，消泡剂选自硅氧烷消泡剂中的一种，表面活性剂选自聚氧乙烯氢化蓖麻油和蔗糖脂肪酸酯中的一种或多种；本发明提供的β‑环糊精‑L‑薄荷醇复合物、L‑薄荷醇水溶液和L‑薄荷醇纳米乳的制备方法简单，且能够缓解ERCP的胆道痉挛。

技术领域

Technical Field

The invention belongs to the technical field of medicines, and particularly relates to an ERCP (intestinal canal spasm) special compound for relieving biliary spasm, a preparation and a preparation method thereof.

背景技术

Background

ERCP (endoscopic retrograde cholangiopancreatography) is one of the main means for minimally invasive treatment of the biliary-pancreatic diseases at present, an endoscope is inserted into a duodenum descending part through the mouth, enters a bile duct or a pancreatic duct through a special duodenal papilla introduction instrument, contrast agent is injected under X-ray fluoroscopy for contrast, and a sub-endoscope/ultrasonic probe is introduced for observation to complete diagnosis of biliary and pancreatic diseases, and a general name of a technology of corresponding interventional therapy is implemented on the basis of diagnosis.

Menthol has antipruritic, analgesic, antiseptic, irritant, anesthetic, refreshing and anti-inflammatory effects, and can be used for treating headache, neuralgia, pruritus, respiratory inflammation, atrophic rhinitis, hoarseness, etc., but menthol has not been reported to be used for relieving ERCP biliary spasm.

发明内容

Disclosure of Invention

In order to solve the technical problems, the invention provides an ERCP (enterocholecystokinin) -specific biliary spasm relieving compound, which comprises L-menthol, beta-cyclodextrin or derivatives thereof in a mass ratio of 1-3.

The research of the invention finds that the compound prepared by the L-menthol and the beta-cyclodextrin can be used for relieving the biliary spasm of the ERCP.

Furthermore, the compound also comprises glycyrrhetinic acid, wherein the mass ratio of L-menthol, beta-cyclodextrin or derivatives thereof to glycyrrhetinic acid is 1-3.

In the research and development process, the L-menthol has the effect of inhibiting the central nervous system, so that when the ERCP is clinically implemented, if the L-menthol is directly used as a spasmolytic, the L-menthol is easy to generate the effect of inhibiting the central nervous system of a patient; however, the invention unexpectedly discovers that after the L-menthol, the beta-cyclodextrin or the derivative thereof and the glycyrrhetinic acid are prepared into the compound, the compound can play a good role in spasmolysis, is convenient for doctors to carry out ERCP operation on patients, can not inhibit the central nervous system, and solves the problem that the L-menthol can not be applied to spasmolysis before ERCP.

Further, the beta-cyclodextrin derivative comprises one or more of hydroxypropyl-beta-cyclodextrin, methylated beta-cyclodextrin, and sulfobutyl ether-beta-cyclodextrin.

Further, the compound consists of L-menthol, beta-cyclodextrin or derivatives thereof and glycyrrhetinic acid in a mass ratio of 1-3 to 13-15, wherein the beta-cyclodextrin and the derivatives thereof are hydroxypropyl-beta-cyclodextrin and sulfobutyl ether-beta-cyclodextrin in a mass ratio of 1-2.

The invention specifically limits that the beta-cyclodextrin and the derivative thereof are hydroxypropyl-beta-cyclodextrin and sulfobutyl ether-beta-cyclodextrin in a mass ratio of 1-2-8-12, and can obviously reduce the dosage of the beta-cyclodextrin and the derivative thereof under the condition of ensuring the same encapsulation efficiency, thereby reducing the dosage of the compound.

The invention also provides an L-menthol nanoemulsion which comprises L-menthol, a surfactant and a defoaming agent, wherein the mass ratio of the L-menthol to the surfactant to the defoaming agent is 1-2; the defoaming agent is selected from one of siloxane defoaming agents, and the surfactant is selected from one or more of polyoxyethylene hydrogenated castor oil and sucrose fatty acid ester.

The invention also provides an L-menthol aqueous solution, which comprises a compound and water, wherein the mass ratio of L-menthol to water is 1-3.

The invention discovers that if the compound is prepared into powder, when the compound is mixed with water and then is guided into the biliary tract through an endoscope during use, the compound can not be uniformly distributed in the water, and a part of the compound is attached to the inner wall of the endoscope, so that more compound is needed to achieve the expected effect; the invention can solve the problem of uneven distribution of the compound in water by preparing the compound into an aqueous solution.

Further, the L-menthol aqueous solution is prepared by the following method:

(1) Uniformly mixing beta-cyclodextrin or derivatives thereof with water according to a specified mass ratio to prepare a beta-cyclodextrin aqueous solution;

(2) Adjusting the pH value of the beta-cyclodextrin aqueous solution to 7.4-7.6, adding glycyrrhetinic acid and L-menthol in a specified mass ratio into the beta-cyclodextrin aqueous solution, heating to 50-60 ℃, stirring under the protection of inert gas at the stirring speed of 30-50r/min for 10-15min, cooling, filtering, subpackaging and capping; sterilizing at 115 deg.C for 20-40min to obtain L-menthol aqueous solution.

The L-menthol aqueous solution prepared by the method has better stability and can be stored for 12 months at normal temperature.

Further, the filtration of step (2) is a 0.22um membrane filtration.

Further, the inert gas is nitrogen.

The L-menthol aqueous solution provided by the invention is used for relieving biliary spasm during ERCP.

The beta-cyclodextrin-L-menthol compound and the L-menthol aqueous solution provided by the invention have simple preparation methods, can relieve the biliary spasm of ERCP, and cannot inhibit the central nervous system.

附图说明null实施方式

Detailed Description

Example 1

This example provides a complex comprising 10g of l-menthol and 130g of methylated beta-cyclodextrin;

methylated beta-cyclodextrin is produced by Suzhou Philippine Biotech limited.

Example 2

This example provides a complex comprising 20g of l-menthol and 180g of methylated beta-cyclodextrin;

methylated beta-cyclodextrin is produced by Suzhou Philippine Biotech limited.

Example 3

This example provides a complex comprising 30g of l-menthol and 250g of methylated beta-cyclodextrin;

methylated beta-cyclodextrin is produced by Suzhou Philippine Biotech limited.

Example 4

This example provides a complex comprising 10g of l-menthol, 150g of β -cyclodextrin or a derivative thereof, and 20g of glycyrrhetinic acid; wherein the beta-cyclodextrin or the derivative thereof is methylated beta-cyclodextrin produced by Suzhou Fulu Biotechnology Co., ltd;

the preparation method of the compound comprises the following steps:

s1: uniformly mixing methylated beta-cyclodextrin with specified mass and water, wherein the mass ratio of the methylated beta-cyclodextrin to the water is 1;

s2: adding glycyrrhetinic acid and L-menthol with specified mass into beta-cyclodextrin water solution, heating to 50 ℃, stirring under the protection of inert gas at the stirring speed of 30r/min for 10min, cooling, filtering, and drying to obtain powder; obtaining L-menthol aqueous solution;

wherein, the filtration in the step (2) is the filtration through a 0.22um membrane;

wherein the inert gas is nitrogen.

Example 5

This example provides a complex comprising 10g of l-menthol, 250g of beta-cyclodextrin or a derivative thereof, and 20g of glycyrrhetinic acid; wherein the beta-cyclodextrin or the derivative thereof is hydroxypropyl-beta-cyclodextrin produced by Suzhou Fulu Biotechnology Co., ltd;

the preparation method of the compound comprises the following steps:

s1: uniformly mixing hydroxypropyl-beta-cyclodextrin with specified mass and water, wherein the mass ratio of hydroxypropyl-beta-cyclodextrin to water is 1;

s2: adding glycyrrhetinic acid and L-menthol with specified mass into beta-cyclodextrin water solution, heating to 50 ℃, stirring under the protection of inert gas at the stirring speed of 30r/min for 10min, cooling, filtering, and drying to obtain powder; obtaining L-menthol aqueous solution;

wherein, the filtration in the step (2) is the filtration through a 0.22um membrane;

wherein the inert gas is nitrogen.

Example 6

This example provides a complex comprising 10g of l-menthol, 250g of beta-cyclodextrin or a derivative thereof, and 20g of glycyrrhetinic acid; wherein the beta-cyclodextrin or the derivative thereof is sulfobutyl ether-beta-cyclodextrin produced by Suzhou Fulu Biotechnology Co., ltd;

the preparation method of the compound comprises the following steps:

s1: uniformly mixing sulfobutyl ether-beta-cyclodextrin with water according to the specified mass ratio of 1;

s2: adding glycyrrhetinic acid and L-menthol with specified mass into beta-cyclodextrin water solution, heating to 50 ℃, stirring under the protection of inert gas at the stirring speed of 30r/min for 10min, cooling, filtering, and drying to obtain powder; obtaining L-menthol aqueous solution;

wherein, the filtration of the step (2) is the filtration through a 0.22um membrane;

wherein the inert gas is nitrogen.

Example 7

This example provides a complex comprising 20g of l-menthol, 130g of beta-cyclodextrin or a derivative thereof, and 30g of glycyrrhetinic acid; wherein the beta-cyclodextrin or the derivative thereof comprises 10g of hydroxypropyl-beta-cyclodextrin and 120g of sulfobutyl ether-beta-cyclodextrin, and the hydroxypropyl-beta-cyclodextrin and the sulfobutyl ether-beta-cyclodextrin are both produced by Suzhou Sulu Biotech, inc.;

the preparation method of the compound comprises the following steps:

s1: uniformly mixing hydroxypropyl-beta-cyclodextrin and sulfobutyl ether-beta-cyclodextrin with specified mass and water, wherein the mass ratio of the total amount of the hydroxypropyl-beta-cyclodextrin and the sulfobutyl ether-beta-cyclodextrin to the water is 1;

s2: adding glycyrrhetinic acid and L-menthol with specified mass into beta-cyclodextrin water solution, heating to 55 ℃, stirring under the protection of inert gas at the stirring speed of 40r/min, cooling, filtering, and drying to powder; obtaining L-menthol aqueous solution;

wherein, the filtration of the step (2) is the filtration through a 0.22um membrane;

wherein the inert gas is nitrogen.

Example 8

This example provides a complex comprising 30g of l-menthol, 150g of β -cyclodextrin or a derivative thereof, and 40g of glycyrrhetinic acid; wherein the beta-cyclodextrin or the derivative thereof comprises 30g of hydroxypropyl-beta-cyclodextrin and 120g of sulfobutyl ether-beta-cyclodextrin, and the hydroxypropyl-beta-cyclodextrin and the sulfobutyl ether-beta-cyclodextrin are both produced by Suzhou Philippine biotechnology, inc.;

the preparation method of the compound comprises the following steps:

s1: uniformly mixing hydroxypropyl-beta-cyclodextrin and sulfobutyl ether-beta-cyclodextrin with water according to the specified mass ratio of the total amount of the hydroxypropyl-beta-cyclodextrin and the sulfobutyl ether-beta-cyclodextrin to the water of 1;

s2: adding glycyrrhetinic acid and L-menthol with specified mass into beta-cyclodextrin water solution, heating to 60 ℃, stirring under the protection of inert gas at the stirring speed of 50r/min, cooling, filtering, and drying to obtain powder; obtaining L-menthol aqueous solution;

wherein, the filtration in the step (2) is the filtration through a 0.22um membrane;

wherein the inert gas is nitrogen.

Example 9

This example provides an aqueous solution of L-menthol, consisting of 10g of L-menthol, 130g of beta-cyclodextrin or a derivative thereof, 20g of glycyrrhetinic acid, and 1000g of water, wherein the beta-cyclodextrin or the derivative thereof comprises 10g of hydroxypropyl-beta-cyclodextrin and 120g of sulfobutyl ether-beta-cyclodextrin, both of which are produced by voluminous biotechnology limited, suzhou;

the preparation method of the aqueous solution comprises the following steps:

(1) Uniformly mixing beta-cyclodextrin or derivatives thereof with water according to a specified mass ratio to prepare a beta-cyclodextrin aqueous solution;

(2) Adjusting the pH value of the beta-cyclodextrin aqueous solution to 7.4, adding glycyrrhetinic acid and L-menthol with specified mass into the beta-cyclodextrin aqueous solution, heating to 50 ℃, stirring under the protection of inert gas at the stirring speed of 30r/min for 10min, cooling, filtering, subpackaging and capping; sterilizing at 115 deg.C for 20min to obtain L-menthol aqueous solution;

wherein, the filtration in the step (2) is the filtration through a 0.22um membrane;

wherein the inert gas is nitrogen.

Example 10

This example provides an aqueous solution of L-menthol consisting of 20g of L-menthol, 140g of beta-cyclodextrin or a derivative thereof, 30g of glycyrrhetinic acid, and 1200g of water, wherein the beta-cyclodextrin or the derivative thereof comprises 28g of hydroxypropyl-beta-cyclodextrin and 112g of sulfobutyl ether-beta-cyclodextrin, both hydroxypropyl-beta-cyclodextrin and sulfobutyl ether-beta-cyclodextrin are produced by schefflera biotechnology limited, suzhou;

the preparation method of the aqueous solution comprises the following steps:

(1) Uniformly mixing beta-cyclodextrin or derivatives thereof with water according to a specified mass ratio to prepare a beta-cyclodextrin aqueous solution;

(2) Adjusting the pH value of the beta-cyclodextrin aqueous solution to 7.5, adding glycyrrhetinic acid and L-menthol with specified mass into the beta-cyclodextrin aqueous solution, heating to 55 ℃, stirring under the protection of inert gas at the stirring speed of 40r/min for 12min, cooling, filtering, subpackaging and capping; sterilizing at 115 deg.C for 30min to obtain L-menthol aqueous solution;

wherein, the filtration of the step (2) is the filtration through a 0.22um membrane;

wherein the inert gas is nitrogen.

Example 11

This example provides an aqueous solution of L-menthol, consisting of 30g of L-menthol, 150g of beta-cyclodextrin or a derivative thereof, 40g of glycyrrhetinic acid, and 1500g of water, wherein the beta-cyclodextrin or the derivative thereof comprises 30g of hydroxypropyl-beta-cyclodextrin and 120g of sulfobutyl ether-beta-cyclodextrin, both of which are produced by schefflera biotechnologies, inc, suzhou;

the preparation method of the aqueous solution comprises the following steps:

(3) Uniformly mixing beta-cyclodextrin or derivatives thereof with water according to a specified mass ratio to prepare a beta-cyclodextrin aqueous solution;

(4) Adjusting the pH value of the beta-cyclodextrin aqueous solution to 7.6, adding glycyrrhetinic acid and L-menthol with specified mass into the beta-cyclodextrin aqueous solution, heating to 60 ℃, stirring under the protection of inert gas at a stirring speed of 50r/min for 15min, cooling, filtering, subpackaging and capping; sterilizing at 115 deg.C for 40min to obtain L-menthol aqueous solution;

wherein, the filtration of the step (2) is the filtration through a 0.22um membrane;

wherein the inert gas is nitrogen.

Example 12

The present embodiment provides an L-menthol nanoemulsion comprising L-menthol, a surfactant, and a defoaming agent, wherein the mass ratio of the L-menthol to the surfactant to the defoaming agent is 1.5; the defoaming agent is polydimethylsiloxane-silicon dioxide mixture fluid, and the surfactant is polyoxyethylene hydrogenated castor oil.

Comparative example 1

This comparative example provides a complex comprising 20g of l-menthol, 130g of beta-cyclodextrin or a derivative thereof and 30g of poria; wherein the beta-cyclodextrin or the derivative thereof comprises 10g of hydroxypropyl-beta-cyclodextrin and 120g of sulfobutyl ether-beta-cyclodextrin, and the hydroxypropyl-beta-cyclodextrin and the sulfobutyl ether-beta-cyclodextrin are both produced by Suzhou Paolu Biotech limited;

the preparation method of the compound comprises the following steps:

s1: uniformly mixing hydroxypropyl-beta-cyclodextrin and sulfobutyl ether-beta-cyclodextrin with specified mass and water, wherein the mass ratio of the total amount of the hydroxypropyl-beta-cyclodextrin and the sulfobutyl ether-beta-cyclodextrin to the water is 1;

s2: adding poria cocos element with specified mass into beta-cyclodextrin water solution, heating to 55 ℃, stirring under the protection of inert gas at the stirring speed of 40r/min, cooling, filtering, and drying to obtain powder; obtaining L-menthol aqueous solution;

wherein, the filtration in the step (2) is the filtration through a 0.22um membrane;

wherein the inert gas is nitrogen.

Comparative example 2

This comparative example provides a complex comprising 20g of L-menthol, 130g of beta-cyclodextrin or a derivative thereof and 30g of strychnine; wherein the beta-cyclodextrin or the derivative thereof comprises 10g of hydroxypropyl-beta-cyclodextrin and 120g of sulfobutyl ether-beta-cyclodextrin, and the hydroxypropyl-beta-cyclodextrin and the sulfobutyl ether-beta-cyclodextrin are both produced by Suzhou Paolu Biotech limited;

the preparation method of the compound comprises the following steps:

s1: uniformly mixing hydroxypropyl-beta-cyclodextrin and sulfobutyl ether-beta-cyclodextrin with water according to the specified mass ratio of the total amount of the hydroxypropyl-beta-cyclodextrin and the sulfobutyl ether-beta-cyclodextrin to the water of 1;

s2: adding a prescribed mass of tannin and L-menthol into a beta-cyclodextrin aqueous solution, heating to 55 ℃, stirring under the protection of inert gas at the stirring speed of 40r/min, cooling, filtering and drying to obtain powder; obtaining L-menthol aqueous solution;

wherein, the filtration of the step (2) is the filtration through a 0.22um membrane;

wherein the inert gas is nitrogen.

Comparative example 3

This comparative example provides a complex comprising 20g turpentine, 130g beta-cyclodextrin or a derivative thereof, and 30g glycyrrhetinic acid; wherein the beta-cyclodextrin or the derivative thereof comprises 10g of hydroxypropyl-beta-cyclodextrin and 120g of sulfobutyl ether-beta-cyclodextrin, and the hydroxypropyl-beta-cyclodextrin and the sulfobutyl ether-beta-cyclodextrin are both produced by Suzhou Paolu Biotech limited;

the preparation method of the compound comprises the following steps:

s1: uniformly mixing hydroxypropyl-beta-cyclodextrin and sulfobutyl ether-beta-cyclodextrin with water according to the specified mass ratio of the total amount of the hydroxypropyl-beta-cyclodextrin and the sulfobutyl ether-beta-cyclodextrin to the water of 1;

s2: adding glycyrrhetinic acid and turpentine oil with specified mass into beta-cyclodextrin water solution, heating to 55 ℃, stirring under the protection of inert gas at the stirring speed of 40r/min, cooling, filtering, and drying to obtain powder; obtaining turpentine water solution;

wherein, the filtration of the step (2) is the filtration through a 0.22um membrane;

wherein the inert gas is nitrogen.

Comparative example 4

This example provides an aqueous solution of L-menthol consisting of 20g of L-menthol, 140g of beta-cyclodextrin or a derivative thereof, 30g of glycyrrhetinic acid, and 1200g of water, wherein the beta-cyclodextrin or the derivative thereof comprises 28g of hydroxypropyl-beta-cyclodextrin and 112g of sulfobutyl ether-beta-cyclodextrin, both hydroxypropyl-beta-cyclodextrin and sulfobutyl ether-beta-cyclodextrin are produced by schefflera biotechnology limited, suzhou;

the preparation method of the aqueous solution comprises the following steps:

(1) Uniformly mixing beta-cyclodextrin or derivatives thereof with water according to a specified mass ratio to prepare a beta-cyclodextrin aqueous solution;

(2) Adjusting the pH value of the beta-cyclodextrin aqueous solution to 7.5, adding glycyrrhetinic acid and L-menthol with specified mass into the beta-cyclodextrin aqueous solution, heating to 55 ℃, then starting stirring at the stirring speed of 40r/min for 12min, cooling, filtering, subpackaging and capping; sterilizing at 115 deg.C for 30min to obtain L-menthol water solution;

wherein, the filtration in the step (2) is the filtration through a 0.22um membrane.

Comparative example 5

This example provides an aqueous solution of L-menthol consisting of 20g of L-menthol, 140g of beta-cyclodextrin or a derivative thereof, 30g of glycyrrhetinic acid, and 1200g of water, wherein the beta-cyclodextrin or the derivative thereof comprises 28g of hydroxypropyl-beta-cyclodextrin and 112g of sulfobutyl ether-beta-cyclodextrin, both hydroxypropyl-beta-cyclodextrin and sulfobutyl ether-beta-cyclodextrin are produced by schefflera biotechnology limited, suzhou;

the preparation method of the aqueous solution comprises the following steps:

(1) Uniformly mixing beta-cyclodextrin or derivatives thereof with water according to a specified mass ratio to prepare a beta-cyclodextrin aqueous solution;

(2) Adding glycyrrhetinic acid and L-menthol with specified mass into beta-cyclodextrin water solution, heating to 55 deg.C, stirring under inert gas protection at stirring speed of 40r/min for 10-15min, cooling, filtering, packaging, and capping; sterilizing at 115 deg.C for 30min to obtain L-menthol water solution;

wherein, the filtration in the step (2) is the filtration through a 0.22um membrane;

wherein the inert gas is nitrogen.

Test example 1 Effect of L-menthol Complex on bile flow and cholecystokinetic Bar in rats

1 grouping

36 rats were divided into test 1-9 groups and control 1-3 groups, each group containing 3 rats.

2 experimental animals: SD rat provided by Chinese food and drug testing institute. Quality certification number of experimental animal, SCXK (Jing) 2017-0005.

3, anesthetic agent: pentobarbital sodium (sodium diethylmalonylurea), a product of Shanghai chemical reagents, china pharmaceutical company, imported for separate packaging, batch number 2016-06-24.

4, animal feeding: tumor hospital experimental animal center, tumor institute of Chinese medical academy of sciences, SPF environment. License number for experimental animals: SYXK (Jing) 2014-0003.

Feed: edible Co ⁶⁰ Sterilized feed, SPF grade, product feed production license number of kyoto australia synergetics feeds ltd: SCXK (Jing) 2005-007.

5, drinking water: sterilizing water, and sterilizing at high temperature and high pressure.

6, pollutants: the levels of contaminants in food and water were below levels that interfered with and affected the results of the study.

And 7, environment: the animal is living in controlled environment with room temperature 22-25 deg.c, daily temperature difference not more than 3 deg.c, humidity 40-70%, 12 hr illumination, alternate light and dark, air cleanliness 100 level, ammonia concentration not more than 14mg/m ³ Noise is less than or equal to 60 decibels, the working illumination is 150-300 lux, and the animal illumination is 100-200 lux.

8 animal quarantine and inspection: is responsible for the experimental animal center of the tumor hospital of Chinese academy of medical sciences.

9 apparatus equipment:

medical purification bench, model CJ-2F, product of von experimental animal facilities ltd, suzhou. An electronic precision balance, a product of Aohaus corporation in America, and the measuring range is 0.01 g-510 g. An electronic digital display caliper is a product of Guilin Guangdong digital measurement and control GmbH, model SF2000, resolution ratio of 0.01mm, specification of 0-150 mm, standard GB/T14899-94, carbon dioxide inhalation anesthesia facility, tension transducer and a product of Shanghai Xinman scientific and education technology Limited.

10. Grouping and administration:

before the experiment, rats in each group are fasted without water prohibition, anesthetized with 0.6% pentobarbital sodium after 16h, as shown in figure 1, after complete anesthesia, the rats are fixed, a small opening is cut along the median line of the abdomen of the rats by a surgical knife, the fundus is found, the duodenum is exposed along the fundus, the duodenum is turned over, a white thick bile duct is found in the mesentery connecting the duodenum with the stomach and the pancreas, two silk threads are penetrated under the bile duct, the duodenum is ligated by the end of the duodenum, a V-shaped incision is made towards the liver direction, a plastic thin tube with the diameter of about 0.6mm is inserted as a drainage tube, light yellow and green bile flows out, the catheter is ligated and fixed, the bile is collected by a 2mL EP tube at the other end, and the abdominal wall is closed by a hemostatic clamp after the operation, covering by using gauze soaked with normal saline, irradiating by using a sodium lamp to keep the body temperature of a rat at 37 +/-0.5 ℃, collecting 30min of bile as bile flow before administration after bile flows out stably for 10min, then locally administering the bile to a gall bladder near common duodenum, respectively administering the compounds of groups 1-5 and 9 in test 1-6, respectively administering the compounds of groups 1-4 in control 1-4, uniformly adjusting the compounds with water before use in each group, wherein the mass ratio of L-menthol to water in the compounds is 150, the administration amount of each group is 300mg/kg, collecting the bile once every 30min after administration for 1 hour, recording the bile flow, and the test result is shown in table 1; after the experiment is finished, the gallbladder of each rat is taken, muscle strips with the length of 6mm and the width of 2mm are prepared from the gallbladder along the longitudinal axis of the gallbladder, a mucous layer and a serosal layer are removed, each gallbladder is made into two muscle strips, the prepared muscle strips are connected to a hook of a tension transducer, the temperature of a perfusion muscle groove is set to be 37 ℃, then freshly prepared Krebs liquid is poured in, the hook gives 1.Og initial tension as a preload, and after the muscle strips shrink regularly and spontaneously with low amplitude and stability for about 10min, acetylcholine (Ach) solution (the concentration of a reaction system is 3.46 multiplied by 10^ is added ^-8 mol/L), smooth muscle contraction, and a sharp increase in tension, which is the tension after Ach, followed by administration of the corresponding drug to each group of rats, administration of the complexes of examples 1-5 to each group of experiments 1-5, and administration of controls 1-4 to each group of controls 1-4The compound is prepared uniformly by water before use, the mass ratio of L-menthol to water in the compound is 1; post-drug tonicity = post-administration tonicity-basic tonicity; the tension was adjusted to about 1.0g before the experiment. After it had stabilized for 10min, the baseline was traced and the tension recorded, which was the base tension, the results are shown in table 2.

11. Data statistics and analysis

Analysis of variance was performed with SPSS 17.0 statistical software, all data are expressed in x + -s, and the treatment between groups of two samples was performed using the t-test. Data are averaged. + -. Standard deviationDenotes, comparison of groups, P<A statistical difference of 0.05 was significant.

TABLE 1 Effect of the complexes on bile flow in SD rats

TABLE 2 Effect of groups of complexes on the gallbladder muscle strips of SD rats

As can be seen from tables 1 and 2, the L-menthol and the methylated β -cyclodextrin are used in the groups 1 to 3 of the tests, which can effectively inhibit the biliary spasm, the glycyrrhetinic acid is added to the compound in the groups 4 to 6 of the tests on the basis of the groups 1 to 3 of the tests, the effect of inhibiting the biliary spasm is equivalent to that of the groups 1 to 3 of the tests, the effect of obviously reducing the tension of the cholecystokinetic muscle strips of SD rats is also equivalent, the effect of inhibiting the biliary spasm of the groups 7 to 9 of the tests is equivalent to that of the groups 4 to 6 of the tests, but the dosage of the β -cyclodextrin or the derivative thereof can be obviously reduced by specifically limiting the type of the β -cyclodextrin or the derivative thereof, the dosage of the raw materials can be saved while the original technical effect of the compound is ensured, the poria cocos hormone or the stilben is respectively used in the groups 1 to 2 of the tests instead of the glycyrrhetinic acid, the tension of the cholecystokinetic muscle strips of SD rats is slightly higher than that in the groups 4 to 6 of the tests, the group 3 of the controls uses the turpentine instead of the L, which has a poor effect of inhibiting the biliary spasm; the results prove that the compound provided by the invention has the effects of reducing the tension of the gallbladder sphincter Oddi and promoting the relaxation of the sphincter, and simultaneously can promote the secretion of bile and relieve the spasm of the cholecystectomy muscle strip caused by acetylcholine.

Test example 2 groups of complexes on rat central nervous system H ³ Inhibitory Effect of labeled 5-HT reuptake

Taking 60 male rats of 150-200g, dividing into 12 groups, each group comprising 5 rats, respectively performing intragastric administration with the compound of examples 1-9 and comparative examples 1-3, wherein the intragastric administration amount is 100mg/kg, 30min after intragastric administration, cutting neck, taking brain, stripping hypothalamus, weighing, adding 0.32M sucrose solution with concentration of 25ml/g, homogenizing in ice bath, freeze-centrifuging the homogenate at the rotation speed of 1000r/min for 15min, taking supernatant, freeze-centrifuging the supernatant at the rotation speed of 15000r/min for 15min, taking precipitate, diluting with 0.32M sucrose solution with concentration of 25ml/g at the ratio of 10ml/g into diluent for standby, and using freshly prepared Krebs solution to dilute H ³ Diluting the marked monoamine transmitter 200 times, mixing with 800 μ l diluent and diluent (25 μ l) prepared from hypothalamus of each group of rats, filtering with whatman GFB filter paper after 5min, washing with Krebs solution, drying with filter paper, pouring into scintillation solution, and measuringDetermining the scintillation frequency (CPM) within one minute, subtracting the CPM value at 0-4 ℃ from the CPM value at 37 ℃ under the same condition to obtain the monoamine transmitter reuptake value of the central nervous system, and detecting the inhibition condition of the drug on the monoamine transmitter reuptake, wherein the result is shown in Table 3.

TABLE 3 inhibition of the central nervous system in rats by various groups of complexes

As can be seen from Table 3, examples 4 to 9 are given for rat central nervous system H ³ The marked 5-HT reuptake inhibition rate is obviously lower than that of examples 1-3, and the marked 5-HT reuptake inhibition rate is proved to be extremely low by using the glycyrrhetinic acid and the L-menthol for compatibility, so that the prepared compound has extremely low inhibition on the central nervous system and is suitable for being used as a spasmolytic before ERCP, while the compound of comparative examples 1-3 replaces the glycyrrhetinic acid or the L-menthol respectively, so that the prepared compound has strong inhibition on the central nervous system and is not suitable for being used as a spasmolytic before ERCP.

Test example 3 encapsulation efficiency test

Taking 2 mu L of menthol standard substance, then respectively taking the compounds of examples 1-5, wherein each group of compounds contains 2 mu L of menthol, placing the menthol standard substance and each group of compounds in a 20ml empty bottle, balancing for 30min at 60 ℃, detecting the peak area of the menthol by using a 2.3.22 chromatographic method, and calculating the encapsulation efficiency, wherein the result is shown in a table 4, and the calculation formula of the encapsulation efficiency is as follows:

encapsulation ratio (%) = (1-area of menthol peak in complex/area of menthol peak in control) × 100%.

TABLE 4 encapsulation efficiency determination test results

As can be seen from table 4, the encapsulation efficiencies of the complexes of examples 5-9 are relatively small, demonstrating that the use of a specific ratio of hydroxypropyl- β -cyclodextrin and sulfobutyl ether- β -cyclodextrin can significantly reduce the amount of β -cyclodextrin used with similar encapsulation efficiencies.

Example 4 stability test

The penicillin bottles of the examples 10-11 and the comparative examples 4-5 are sealed according to the conventional sealing method of the penicillin bottles for injection, the penicillin bottles are placed under the conditions of 4 ℃ plus or minus 2 ℃ and 60% plus or minus 10% of relative humidity for long-term storage, 5 bottles of each group of aqueous solution are sampled, the labeled content of the L-menthol in the aqueous solution is measured, the results are averaged, and the test results are shown in the table 5.

TABLE 5 stability test results for each aqueous solution group

As can be seen from table 5, the stability of examples 10 to 11 is significantly higher than that of comparative examples 4 to 5, which demonstrates that the method for preparing an aqueous solution according to the present invention can significantly improve the stability of aqueous solution, and when some steps in the method are modified or deleted, the stability of the aqueous solution can be significantly reduced.

Finally, it should be noted that the above-mentioned embodiments are only for illustrating the technical solutions of the present invention and not for limiting, although the present invention is described in detail with reference to the preferred embodiments, it should be understood by those skilled in the art that modifications or equivalent substitutions may be made on the technical solutions of the present invention without departing from the spirit and scope of the technical solutions of the present invention, and all of them should be covered by the claims of the present invention.