CN108078929B

CN108078929B - Preparation method of bupivacaine multivesicular liposome and bupivacaine multivesicular liposome preparation

Info

Publication number: CN108078929B
Application number: CN201711498422.6A
Authority: CN
Inventors: 王秋云
Original assignee: Guangzhou Bositao Controlled Release Pharmaceutical Co ltd
Current assignee: Guangzhou Bositao Controlled Release Pharmaceutical Co ltd
Priority date: 2017-12-28
Filing date: 2017-12-28
Publication date: 2021-01-01
Anticipated expiration: 2037-12-28
Also published as: CN108078929A

Abstract

The invention discloses a preparation method of bupivacaine multivesicular liposome and a bupivacaine multivesicular liposome preparation, and relates to the field of bupivacaine medicinal preparations. The preparation method of the bupivacaine multivesicular liposome comprises the following steps: to a first emulsion formed from an oil phase containing an organic solvent and a first aqueous phase containing bupivacaine, a first solution for forming a double emulsion is obtained by adding neutral lipids. Compared with the existing preparation method of the bupivacaine multivesicular liposome, the preparation method provided by the invention has the advantages that the step of adding neutral lipid after the primary emulsion is formed by the oil phase containing the organic solvent and the first water phase containing bupivacaine can overcome the problem of breaking the multivesicular liposome caused by the subsequent step of removing the organic solvent, so that the prepared multivesicular liposome is round in shape, basically contains no lipid fragments, and the long-acting slow-release effect of the bupivacaine medicament is improved.

Description

Preparation method of bupivacaine multivesicular liposome and bupivacaine multivesicular liposome preparation

Technical Field

The invention relates to the field of bupivacaine medicinal preparations, in particular to a preparation method of a bupivacaine multivesicular liposome and a bupivacaine multivesicular liposome preparation.

Background

Bupivacaine is a BCS 1 medicament, has good water solubility and short half-life, and is usually developed into a multivesicular liposome preparation in order to enable an injection to achieve the effect of long-acting and slow release.

The microstructure of multivesicular liposomes (MVLs) is the polymerization of a plurality of liposomes into a sphere, each liposome consists of a hydrophobic membrane and an internal aqueous phase, and the drug is dissolved in the internal aqueous phase. The multivesicular liposome is dispersed in an aqueous phase to form an injection. The multivesicular liposome is a non-concentric alveolate liposome, a plurality of large vesicles separated by lipid bilayers are arranged in the multivesicular liposome, the unique structure endows the liposome with stronger rigidity, when one vesicle is ruptured, the drug is only released from the ruptured vesicle, and the complete vesicle can still keep the original shape, so that the leakage rate of the drug is reduced, and the drug is not suddenly released when being released from the liposome, thereby realizing the control of the slow release of the drug in the time range of one day to several weeks.

However, the bupivacaine-containing multivesicular liposomes prepared by the conventional method had ruptured multivesicular liposomes and ruptured phospholipid fragments, and the particle size of the sample was reduced. The long-acting slow release effect of the multivesicular liposome mainly depends on non-concentric alveolates, and the multivesicular liposome is gradually broken from outside to inside during release to release the medicine. The broken multivesicular liposome can cause the drug not to reach the expected long-acting slow-release effect, even the phenomenon of burst release and the like, and the treatment effect of the drug is influenced.

In view of this, the invention is particularly proposed.

Disclosure of Invention

The invention aims to provide a preparation method of a bupivacaine multivesicular liposome, which can greatly reduce the rupture quantity of the bupivacaine multivesicular liposome and even realize the effect of the unbroken bupivacaine multivesicular liposome.

Another object of the present invention is to provide a bupivacaine multivesicular liposome preparation.

The invention is realized by the following steps:

a preparation method of bupivacaine multivesicular liposome, which comprises the following steps: to colostrum formed from an oil phase containing an organic solvent and a first aqueous phase containing bupivacaine (i.e. W/O water-in-oil colostrum) is added neutral lipids to obtain a first solution for forming a multiple emulsion.

Further, in some embodiments of the present invention, the neutral lipid is Tricaprylin (TC).

Neutral lipids such as TC, which have only hydrophobic ends and no hydrophilic ends, are key materials for the association of multiple liposomes into multivesicular liposomes.

Further, in some embodiments of the invention, after the addition of neutral lipids to the colostrum, the preparation method further comprises a re-emulsion forming step:

the multiple emulsion forming step comprises: and stirring the first solution at a preset temperature which is higher than or equal to 41 ℃.

However, the inventors have further found that when the heating temperature is too high, the organic solvent in the oil phase is volatilized, and the colostrum is reduced in fluidity and hardly transferred.

Further, in some embodiments of the invention, the preset temperature is 41 to 55 ℃.

Further, in some embodiments of the invention, the speed of agitation is less than 5000rpm and the time of agitation is from 1 to 5 min.

Further, in some embodiments of the invention, the speed of agitation is 2000-.

The stirring speed is controlled at 2000-5000rpm, and the stirring time is controlled within 1-5min, so that the neutral lipid can be uniformly dispersed in the oil phase, the structural integrity of the primary emulsion is kept, and the damage of the stirring to the structure of the primary emulsion is avoided.

Further, in some embodiments of the invention, the multiple emulsion forming step further comprises:

and mixing the first solution with the second water to obtain a second solution for forming the multiple emulsion.

Further, in some embodiments of the invention, the second aqueous phase comprises: 30-35mg/mL glucose and 8-12mM lysine.

shearing the second solution to form a multiple emulsion (i.e. a W/O/W water-in-oil-in-water multiple emulsion containing bupivacaine).

Further, in some embodiments of the invention, the second solution is sheared at 3500-4500rpm for 15-30 seconds.

Further, in some embodiments of the present invention, after the multiple emulsion forming step, the preparation method further comprises: removing the organic solvent;

the organic solvent removing step comprises: mixing the double emulsion with inert gas to remove the organic solvent in the double emulsion.

For example, the organic solvent is removed by atomizing the mixture of inert gas flow and double emulsion fluid flow, or the organic solvent is removed by filling the inert gas into the double emulsion.

Further, in some embodiments of the invention, the inert gas is nitrogen.

Further, in some embodiments of the invention, the mass ratio of the added neutral lipid to bupivacaine in the colostrum is 3 (80-120).

The amount of neutral lipid such as TC may affect the sustained-release effect of the drug within a certain range, and an increase in the amount of neutral lipid may cause: firstly, the particle size of the multivesicular liposome is increased, but the multivesicular liposome can be broken due to the final step of controlling the particle size of the product; ② the multivesicular liposome has stronger rigidity, which affects the slow release effect of the drug and the curative effect.

If the amount of the neutral fat is excessively large or small, the production effect may be affected, for example, the milk may not be formed, the single sac may be formed, and the like.

Further, in some embodiments of the present invention, the solvent of the first aqueous phase is water, and the first aqueous phase further comprises one or a combination of the following solutes: glucuronic acid, hydrochloric acid and phosphoric acid;

the solvent of the oil phase is organic solvent, and one or a combination of a plurality of solutes contained in the oil phase: erucyl lecithin (DEPC), dipalmitoyl phosphatidylglycerol (DPPG), and cholesterol.

Wherein DEPC and DPPG are amphiphatic, i.e. phospholipid molecules, have hydrophilic groups and hydrophobic groups, and are main film-forming materials.

Further, in some embodiments of the invention, the first aqueous phase contains 50-65mg/mL bupivacaine, 140-160mM glucuronic acid, 13-17mM hydrochloric acid, and 18-22mM phosphoric acid.

The oil phase contains: 16.5-19.5mM DEPC, 3.5-4.8mM DPPG, and 28-32mM cholesterol.

Further, in some embodiments of the invention, the method of preparation further comprises a colostrum-forming step prior to adding neutral lipids to the primary milk;

the colostrum forming step comprises: the first aqueous phase and the oil phase were mixed and placed in an ice bath (0-10 ℃) for shearing.

Shearing the mixed solution of the first water phase and the oil phase under the ice bath condition, reducing the temperature of the solution and avoiding the volatilization of the organic solvent in the oil phase.

Further, in some embodiments of the invention, the conditions of shearing are: the rotation speed of 15000 and 17000rpm and the time of 7-12 min.

Further, in some embodiments of the invention, the volume ratio of the oil phase to the first organic phase is 1 (2-8).

The inventors have found that if the ratio of the first aqueous phase to the oil phase is 1:1, the colostrum has poor fluidity after formation, which is not favorable for achieving dispersibility of the subsequently added TC, increasing the volume of the oil phase, and making the TC uniformly dispersible. However, the addition of the oil phase volume and the use amount are too large, which can affect two links:

(1) a larger amount of organic solvent needs to be removed before the compound emulsion is formed, energy is lost, and the cost is increased;

(2) if the organic solvent is not removed before the formation of the multiple emulsion, the formation effect of the multiple emulsion can be influenced, multivesicular liposome is difficult to form, single-vesicular liposome tends to be formed, or a large amount of organic solvent is taken away during the removal of the organic solvent, so that the liposome is broken and the fragments are increased.

Further, in some embodiments of the invention, the organic solvent is chloroform or dichloromethane.

A bupivacaine multivesicular liposome preparation, which is prepared by the preparation method. The bupivacaine multivesicular liposome preparation contains less or even no broken multivesicular liposomes.

The invention also aims to provide a bupivacaine multivesicular liposome preparation device.

The invention has the following beneficial effects:

compared with the existing preparation method of the bupivacaine multivesicular liposome, the preparation method of the bupivacaine multivesicular liposome provided by the invention comprises the step of adding neutral lipid into primary emulsion formed by an oil phase containing an organic solvent and a first water phase containing bupivacaine to obtain a first solution for forming multiple emulsion; the preparation method can overcome the problem of breaking the multivesicular liposome caused by the subsequent step of removing the organic solvent by adding the neutral lipid into the colostrum after the formation of the colostrum, so that the prepared multivesicular liposome has round shape, basically contains no phospholipid fragments, and improves the long-acting slow-release effect of the bupivacaine medicament.

In addition, the bupivacaine multivesicular liposome preparation device provided by the invention can be used for firstly forming primary emulsion, then adding neutral lipid and mixing to obtain a first solution for forming multiple emulsion, and can be used for preparing multivesicular liposome which does not contain phospholipid fragments and has a round shape.

Drawings

In order to more clearly illustrate the technical solutions of the embodiments of the present invention, the drawings needed to be used in the embodiments will be briefly described below, it should be understood that the following drawings only illustrate some embodiments of the present invention and therefore should not be considered as limiting the scope, and for those skilled in the art, other related drawings can be obtained according to the drawings without inventive efforts.

Fig. 1 is a morphological observation view under a microscope of a multiple emulsion containing bupivacaine multivesicular liposomes prepared by the method for preparing bupivacaine multivesicular liposomes provided by the embodiment 1 of the invention before removing an organic solvent;

fig. 2 is a morphological observation image under a microscope of a multiple emulsion containing bupivacaine multivesicular liposomes prepared by the method for preparing bupivacaine multivesicular liposomes provided in example 1 of the present invention after removing an organic solvent;

FIG. 3 is a morphological observation under a microscope of a multiple emulsion containing bupivacaine multivesicular liposomes prepared by the method for preparing bupivacaine multivesicular liposomes provided by comparative example 1 of the present invention before removing an organic solvent;

FIG. 4 is a morphological observation image under a microscope of a multiple emulsion containing a bupivacaine multivesicular liposome prepared by the method for preparing the bupivacaine multivesicular liposome provided by the comparative example 1 of the invention after removing an organic solvent;

FIG. 5 is a schematic structural diagram of a bupivacaine multivesicular liposome preparation apparatus provided in example 2 of the present invention;

fig. 6 is a schematic view of an internal structure of a three-way device provided in embodiment 2 of the present invention;

fig. 7 is a schematic view of an internal structure of a three-way device provided in embodiment 2 of the present invention, in which a screen mesh is installed;

fig. 8 is a morphological observation view under a microscope of a multiple emulsion containing bupivacaine multivesicular liposomes prepared by the method for preparing bupivacaine multivesicular liposomes provided in example 3 of the present invention after removing the organic solvent;

fig. 9 is a morphological observation view under a microscope of a multiple emulsion containing bupivacaine multivesicular liposomes prepared by the method for preparing bupivacaine multivesicular liposomes provided in example 4 of the present invention after removing the organic solvent;

fig. 10 is a morphological observation view under a microscope of a multiple emulsion containing bupivacaine multivesicular liposomes prepared by the method for preparing bupivacaine multivesicular liposomes provided in example 5 of the present invention after removing the organic solvent;

fig. 11 is a morphological observation view under a microscope of a multiple emulsion containing bupivacaine multivesicular liposomes prepared by the method for preparing bupivacaine multivesicular liposomes provided in example 6 of the present invention after removing the organic solvent;

fig. 12 is a morphological observation view under a microscope of a multiple emulsion containing bupivacaine multivesicular liposomes prepared by the method for preparing bupivacaine multivesicular liposomes provided in example 7 of the present invention after removing the organic solvent;

fig. 13 is a morphological observation view under a microscope of a multiple emulsion containing bupivacaine multivesicular liposomes prepared by the method for preparing bupivacaine multivesicular liposomes provided in example 8 of the present invention after removing the organic solvent.

Icon: 1-a first aqueous phase storage tank, 2-an oil phase storage tank, 3-a neutral fat storage tank, 4-a first aqueous phase storage tank control valve, 5-an oil phase storage tank control valve, 6-a neutral fat storage tank control valve, 7-a temperature controller, 8-a colostrum reaction tank, 9-a colostrum flow rate control valve, 10-a second aqueous phase storage tank, 11-a second aqueous phase storage tank control valve, 12-a multiple emulsion reaction tank, 13-a multiple emulsion flow rate control valve, 14-a three-way device, 15-a nitrogen generator, 16-a nitrogen control valve, 17-a third aqueous phase storage tank, 18-a third aqueous phase storage tank control valve, 19-a multivesicular liposome receiving tank, and 121-a multiple emulsion inlet pipeline; 141-liquid outlet end, 151-nitrogen gas inlet pipe.

Detailed Description

In order to make the objects, technical solutions and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention will be clearly and completely described below. The examples, in which specific conditions are not specified, were conducted under conventional conditions or conditions recommended by the manufacturer. The reagents or instruments used are not indicated by the manufacturer, and are all conventional products available commercially.

The features and properties of the present invention are described in further detail below with reference to examples.

Example 1

The preparation method of the bupivacaine multivesicular liposome provided by the embodiment comprises the following steps:

1 formation of Water-in-oil colostrum (W/O)

5mL of the first aqueous phase and 25mL of the oil phase were mixed (volume of the first aqueous phase and the oil phase was 1:5), placed in an ice bath and sheared at 16000rpm for 9min to form a water-in-oil colostrum.

Wherein the composition of the first aqueous phase is as follows:

the solvent is water;

the solute is: 60mg/mL bupivacaine, 150mM glucuronic acid, 15mM hydrochloric acid and 20mM phosphoric acid.

The composition of the oil phase was as follows:

the organic solvent is dichloromethane;

the solute is: 18.6mM DEPC, 4.2mM DPPG, and 30mM cholesterol.

2 adding neutral fat (TC)

Adding TC 9mg (mass ratio of TC to bupivacaine in colostrum is 3:100) into colostrum obtained by the above steps, heating colostrum to 41 deg.C, and stirring at a speed of 4500rpm or below for 4 min.

3 forming water-in-oil-in-water double emulsion (W/O/W)

3.1 transfer the colostrum treated in step 2 to a second 25mL aqueous phase and shear at 4000rpm for 20s to form a multiple emulsion containing bupivacaine multivesicular liposomes.

Wherein the composition of the second aqueous phase is as follows:

the solvent is water;

the solute is: 32mg/mL glucose and 10mM lysine solution.

The sampling was performed to determine the particle size and morphology of the multivesicular liposomes, and the results are shown in Table 1 and FIG. 1.

3.2 removal of organic solvents

Nitrogen was mixed with the above double emulsion to remove the organic solvent methylene chloride in the double emulsion.

Sampling was performed to examine the particle size change and morphology of the multivesicular liposomes, and the results are shown in Table 1 and FIG. 2.

3.3 centrifugal concentration (optional step)

Centrifuging the multiple emulsion with organic solvent removed at 200 × g for 10min, and washing the precipitate with 0.9% sodium chloride solution for 3 times to obtain a solution containing bupivacaine multivesicular liposome.

Of course, in some embodiments, the multiple emulsion after removing the organic solvent can also be directly mixed with the third water to obtain the dispersion solvent of the bupivacaine multivesicular liposome.

The composition of the third aqueous phase is the same as the second aqueous phase.

Comparative example 1

The procedure for the preparation of bupivacaine multivesicular liposomes provided in this comparative example is essentially the same as in example 1, except that in this comparative example 9mg of TC is included in the oil phase, i.e. TC is understood to have been added first during the formation of colostrum or used before colostrum formation, and this comparative example 1 does not include step 2 of example 1.

The particle size change of the bupivacaine multivesicular liposome obtained by the preparation method of comparative example 1 before and after the removal of the organic solvent is shown in table 1, and the observation results of the multivesicular lipid morphology before and after the removal of the organic solvent are shown in fig. 3 and 4.

TABLE 1

In table 1, Span is Span, and Span is (D90-D10)/D50, and reflects the degree of width of the normal distribution of particle size, and the smaller Span, the more uniform and narrower the particle size distribution.

As can be seen from the results in Table 1, the multivesicular liposome of comparative example 1 has a significantly reduced particle size after the removal of the organic solvent, while the bupivacaine multivesicular liposome of example 1 has no significant change in particle size before and after the removal of the organic solvent;

and fig. 1 and 2 show the morphology results of the bupivacaine multivesicular liposome of example 1 observed under a microscope, and it can be seen that the bupivacaine multivesicular liposome of example 1 is intact and has no damage before and after the organic solvent is removed; whereas the bupivacaine multivesicular liposomes of comparative example 1 were intact and unbroken (as shown in figure 3) before removal of the organic solvent, but broke (as shown by the arrows in figure 4) after removal of the organic solvent. Example 2

As shown in fig. 5, this example provides a bupivacaine multivesicular liposome preparation apparatus suitable for the preparation method of bupivacaine multivesicular liposome described in example 1, which includes:

the system comprises a first water phase storage tank 1, an oil phase storage tank 2, a neutral fat storage tank 3, a first water phase storage tank control valve 4, an oil phase storage tank control valve 5, a neutral fat storage tank control valve 6, a temperature controller 7, a colostrum reaction tank 8, a colostrum flow rate control valve 9, a second water phase storage tank 10, a second water phase storage tank control valve 11, a multiple-emulsion reaction tank 12, a multiple-emulsion flow rate control valve 13, a tee joint device 14, a nitrogen generator 15, a nitrogen control valve 16, a third water phase storage tank 17, a third water phase storage tank control valve 18 and a multi-vesicle liposome receiving tank 19.

Wherein, the colostrum reaction tank 8 is used for receiving a first mixed solution formed by mixing a first water phase and an oil phase and forming the first mixed solution into colostrum.

And the neutral fat storage tank 3 is used for storing neutral fat, and a neutral fat storage tank control valve 6 is arranged on a conveying pipeline for communicating the neutral fat storage tank 3 with the primary emulsion reaction tank 8 and is used for controlling the conveying of the neutral fat.

A temperature controller 7, which is arranged in the primary emulsion reaction tank 8 and is used for controlling the liquid in the primary emulsion reaction tank 8; the temperature controller 7 may be used to control the temperature at which colostrum is formed from the first mixed solution (e.g. 0-10 ℃) and to heat the colostrum after the neutral fat has been added to it, e.g. to above 41 ℃.

A first aqueous phase storage tank 1 and an oil phase storage tank 2 for storing a first aqueous phase and an oil phase, respectively. Of course, a first water phase storage tank control valve 4 is arranged on a conveying pipeline for communicating the first water phase storage tank 1 with the primary emulsion reaction tank 8 and is used for controlling the conveying of the first water phase; an oil storage tank control valve 5 is arranged on a conveying pipeline for communicating the oil phase storage tank 2 with the colostrum reaction tank 8 and is used for controlling the conveying of the oil phase.

The oil storage tank control valve, the first aqueous phase storage tank control valve and the neutral fat storage tank control valve operate independently of each other.

A mixing line for merging the first aqueous phase and the oil phase is provided at the end of the first aqueous phase transfer line and the end of the oil phase transfer line, that is, the end near the colostrum reaction tank 8, and the mixing line allows the first mixed solution formed by merging the first aqueous phase and the oil phase to be introduced into the colostrum reaction tank 8. The mixing pipeline and the conveying pipeline for conveying neutral fat are independent or separated or do not interfere.

Of course, in other embodiments, the first aqueous phase and the oil phase may be separately introduced into the colostrum reaction tank 8 and then mixed.

A multiple emulsion reaction tank 12 for receiving a second mixed solution formed by the colostrum from the colostrum reaction tank 8 and the second aqueous phase from the second aqueous phase storage tank 10 and forming the second mixed solution into multiple emulsion.

A primary emulsion flow rate control valve 9 for controlling the primary emulsion flow rate is arranged on a conveying pipeline communicated with the primary emulsion reaction tank 8 and the multiple emulsion reaction tank 12; a second water phase storage tank control valve is arranged on a conveying pipeline of the second water phase storage tank 10 communicated with the multiple emulsion reaction tank 12.

And a tee joint device 14 for mixing the nitrogen from the nitrogen generator 15 with the multiple emulsion from the multiple emulsion reaction tank 12 to remove the organic solvent in the multiple emulsion.

The internal structure of the three-way device 14 is shown in fig. 6, the three-way device 14 has a nitrogen inlet pipeline 151 and a multiple emulsion inlet pipeline 121, and the multiple emulsion inlet pipeline 121 wraps the nitrogen inlet pipeline 151. The nitrogen in the nitrogen inlet pipe 151 is mixed with the multiple emulsion in the multiple emulsion inlet pipe 121 at the outlet end 141 near the three-way device 14 to form bupivacaine multivesicular liposome, and the mixed solution flows to the multivesicular liposome receiving tank 19 through the outlet end 141.

In other embodiments, the outlet end 141 of the three-way device 14 is provided with a mesh 142 (as shown in fig. 7) for controlling and/or homogenizing the size of the bupivacaine multivesicular liposomes.

Wherein, a nitrogen control valve 16 is arranged on a delivery pipeline of the nitrogen generator 15 communicated with the three-way device 14.

A composite milk flow rate control valve 13 for controlling the composite milk flow rate is arranged on a conveying pipeline of the composite milk reaction tank 12 communicated with the three-way device 14.

And the multivesicular liposome receiving tank 19 is used for receiving the multiple emulsion after the organic solvent is removed from the three-way device 14.

A third aqueous phase storage tank 17 for storing a third aqueous phase; a third aqueous phase storage tank control valve 18 is arranged on a conveying pipeline communicated with the third aqueous phase storage tank 17 and used for controlling the conveying of the third aqueous phase.

The third aqueous phase can be mixed with the multiple emulsion after removing the organic solvent 151 by controlling the valve 18 of the third aqueous phase storage tank to form the dispersion solvent of the bupivacaine multivesicular liposome.

The composition of the third aqueous phase was the same as that of the second aqueous phase in example 1.

The device for preparing the bupivacaine multivesicular liposome provided by the embodiment can realize the purpose of firstly forming primary emulsion and then adding neutral fat to mix to form multiple emulsion, and further prepare the dispersing solvent of the bupivacaine multivesicular liposome containing less or even not containing the crushed bupivacaine multivesicular liposome.

Example 3

The preparation method of bupivacaine multivesicular liposome provided in this example is basically the same as that of example 1, except that the organic solvent of the oil phase is chloroform in this example. The morphology observation result of the bupivacaine multivesicular liposome prepared in this example under a microscope is shown in fig. 8.

The results in FIG. 8 show that multivesicular liposomes prepared in this example do not contain phospholipid fragments and are round in shape.

Example 4

The preparation method of bupivacaine multivesicular liposomes provided in this example is substantially the same as that of example 1, except that in step 2, colostrum is heated to 55 ℃. The morphology observation result of the bupivacaine multivesicular liposome prepared in this example under a microscope is shown in fig. 9.

The results in FIG. 9 show that multivesicular liposomes prepared in this example do not contain phospholipid fragments and are round in shape.

Example 5

The preparation method of the bupivacaine multivesicular liposome provided in this example is basically the same as that of example 1, except that in this example, the volume ratio of the first aqueous phase to the oil phase is 1: 8.

The morphology observation result of the bupivacaine multivesicular liposome prepared in this example under a microscope is shown in fig. 10.

The results in FIG. 10 show that multivesicular liposomes prepared in this example do not contain phospholipid fragments and are round in shape.

Example 6

The preparation method of the bupivacaine multivesicular liposome provided in this example is basically the same as that of example 1, except that in this example, the volume ratio of the first aqueous phase to the oil phase is 1: 2.

The morphology observation result of the bupivacaine multivesicular liposome prepared in this example under a microscope is shown in fig. 11.

The results in FIG. 11 show that multivesicular liposomes prepared in this example do not contain phospholipid fragments and are round in shape.

Example 7

The preparation method of the bupivacaine multivesicular liposome provided in this example is basically the same as that of example 1, except that, in step 2, the mass ratio of (TC addition to bupivacaine in colostrum) is 3: 120.

The morphology observation result of the bupivacaine multivesicular liposome prepared in this example under a microscope is shown in fig. 12.

The results in FIG. 12 show that multivesicular liposomes prepared in this example do not contain phospholipid fragments and are round in morphology.

Example 8

The preparation method of the bupivacaine multivesicular liposome provided in this example is basically the same as that of example 1, except that, in step 2, the mass ratio of (TC addition to bupivacaine in colostrum) is 3: 80.

The morphology observation result of the bupivacaine multivesicular liposome prepared in this example under a microscope is shown in fig. 13.

The results in FIG. 13 show that multivesicular liposomes prepared in this example do not contain phospholipid fragments and are round in morphology.

In summary, the preparation method of the bupivacaine multivesicular liposome provided by the embodiment of the invention can overcome the problem of breaking of multivesicular liposome caused by the subsequent step of removing organic solvent by adding neutral lipid into colostrum after forming colostrum, so that the prepared multivesicular liposome is round in shape, basically has no phospholipid fragments, and improves the long-acting slow-release effect of bupivacaine medicine.

The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention, and various modifications and changes may be made by those skilled in the art. Any modification, equivalent replacement, or improvement made within the spirit and principle of the present invention should be included in the protection scope of the present invention.

Claims

1. A preparation method of bupivacaine multivesicular liposome is characterized by comprising the following steps: adding neutral lipid to a first emulsion formed from an oil phase containing an organic solvent and a first aqueous phase containing bupivacaine to obtain a first solution for forming a double emulsion; the mass ratio of the added neutral lipid to the bupivacaine in the colostrum is 3 (80-120);

after adding the neutral lipid to the colostrum, the preparation method further comprises a re-emulsion forming step: the multiple emulsion forming step comprises: stirring the first solution at a preset temperature, wherein the preset temperature is 41-55 ℃; the stirring speed is 2000-5000rpm, and the stirring time is 1-5 min;

the method of preparation further comprises a colostrum-forming step prior to adding the neutral lipid to the colostrum; the colostrum forming step comprises: mixing the first water phase and the oil phase, and placing the mixture in an ice bath for shearing; the volume ratio of the first aqueous phase to the oil phase is 1: (2-8).

2. The method for preparing bupivacaine multivesicular liposomes according to claim 1, wherein the multiple emulsion forming step further comprises:

and mixing the stirred first solution with a second water to obtain a second solution for forming the multiple emulsion.

3. The method for preparing bupivacaine multivesicular liposomes according to claim 1, wherein the solvent of the first aqueous phase is water, and the first aqueous phase further comprises one or a combination of several of the following solutes: glucuronic acid, hydrochloric acid and phosphoric acid;

the solvent of the oil phase is an organic solvent, and the oil phase contains one or a combination of more of the following solutes: erucyl lecithin, dipalmitoyl phosphatidylglycerol, and cholesterol.

4. A bupivacaine multivesicular liposome preparation, which is prepared by the method for preparing a bupivacaine multivesicular liposome according to any one of claims 1 to 3.