CN109985075B - Ginkgo leaf extract injection and preparation method thereof - Google Patents

Abstract

The invention discloses a ginkgo leaf extract injection, which is microemulsion of a ginkgo leaf extract, wherein the content of the ginkgo leaf extract is 2.5-4.5 mg/mL. In addition, the preparation method of the injection is also disclosed. The injection has high encapsulation rate of ginkgolide B and bilobalide, has content higher than that of the injection on the market, and has excellent dispersion stability and chemical stability.

Description

Ginkgo leaf extract injection and preparation method thereof

Technical Field

The invention belongs to the technical field of medicine; relates to a ginkgo biloba extract pharmaceutical preparation, in particular to a ginkgo biloba extract injection and a preparation method thereof.

Background

Semen Ginkgo is a tree of Ginkgo of Ginkgoaceae, and is also called semen Ginkgo tree and Duck's foot tree. The leaves, nuts and testa of Ginkgo have corresponding pharmacological action, and the medicinal value of the leaves is the highest. From Song dynasty, China begins to use ginkgo leaves as the medicine, and the ginkgo leaves are dry leaves of ginkgo, are neutral in nature, sweet, bitter and astringent in taste, enter heart and lung channels, and have the effects of astringing lung, relieving asthma, promoting blood circulation and removing blood stasis and the like.

In 1965, German scientists firstly extracted with acetone aqueous solution and then obtained folium Ginkgo extract by 15 procedures of degreasing, removing impurities and enriching. The folium Ginkgo extract is light brown yellow to brown powder, and contains flavonoids (including monoflavone, biflavone and catechin), terpene lactone compounds (including bilobalide A-C and J-M and bilobalide) and ginkgolic acid as main ingredients, and other ingredients; the active ingredients mainly comprise various monoflavonoid glycoside compounds (mainly quercetin, kaempferide and isorhamnetin) and ginkgolide B. The EGB761 extract as standard extract contains total flavonoid glycoside not less than 24%, total lactone not less than 6%, and ginkgolic acid not more than 5 ppm.

Pharmacological research shows that the ginkgo leaf extract has the function of eliminating free radicals, is mainly used for treating circulatory disturbance of brain and peripheral blood flow clinically, and has the protection function on myocardial cells of ischemia-reperfusion injury and diabetic cardiomyopathy. Furthermore, the ginkgo biloba extract can enhance the synthesis of the degreased lipoprotein E of the glial cells, reduce the apoptosis speed of the damaged hippocampal neuronal cells, and relieve the damage of hippocampal tissues, thereby preventing the cerebral apoplexy patients from developing vascular dementia. In addition, the ginkgo biloba extract can better inhibit the over-expression of MMP-2mRNA, and simultaneously has good prevention and treatment effects on pulmonary interstitial fibrosis and lung injury. The research also shows that the ginkgo biloba extract has the function of promoting the mucosa repair and ulcer healing of peptic ulcer patients clinically. Besides the pharmacological actions, the ginkgo leaf extract also has the effects of resisting tumors, reducing serum cholesterol, resisting inflammation, resisting viruses and the like.

The pharmacological active components of folium Ginkgo extract are bilobalide B and bilobalide. However, these two active ingredients have poor solubility and are poorly soluble in water; in addition, both are easily decomposed by heating, and are not suitable for paper cup injection according to the conventional process. The ginkgo biloba injection such as the Shuxuening injection and the gold multi-injection which are on the market at present need to be sterilized at high temperature in the production process, so that the ginkgolide B and the bilobalide are mostly damaged, and the contents of two active ingredients in the injection are lower; meanwhile, the stability of the injection is poor,

in order to solve the problems, Chinese patent CN1267096C discloses a ginkgo leaf extract injection, which comprises 5-99% of ginkgo leaf extract, 1-95% of poloxamer 188 and an alkaline pH value regulator, and the method comprises the steps of firstly injecting poloxamer solution into an aqueous solvent, then adding the ginkgo leaf extract and stirring, then adding the alkaline pH value regulator to regulate the mixture to a physiologically acceptable range, and obtaining the freeze-dried powder injection after vacuum freeze-drying and sealing.

Chinese patent CN1277903C discloses a ginkgolide injection comprising ginkgolide, meglumine, sodium chloride and 10% citric acid solution. The ginkgolide injection product meets the requirement of fingerprint, has controllable product quality, and has good solubilizing effect on ginkgolide under the condition of adding meglumine.

However, the addition of either a basic pH adjuster or meglumine results in ring opening of ginkgolide B and bilobalide. Even if the ring closure reduction is carried out under the acidic condition, the optical activity of the two can be changed, even the structure of the two is transformed, and the pharmacological activity of the two is reduced and even disappears.

Therefore, in view of the above defects in the prior art, there is an urgent need to further find an extract injection of ginkgo biloba leaves and a preparation method thereof, wherein the extract injection can effectively increase the contents of ginkgolide B and bilobalide and can actually improve the stability of ginkgolide B and bilobalide.

Disclosure of Invention

The invention aims to overcome the defects of the prior art and provide the ginkgo biloba extract injection which can effectively improve the contents of ginkgolide B and bilobalide and can also practically improve the stability of the ginkgolide B and the bilobalide and the preparation method thereof.

In order to achieve the above objects, in one aspect, the present invention provides an injection of ginkgo biloba extract, wherein the injection is a microemulsion of ginkgo biloba extract, and the content of the ginkgo biloba extract is 2.5-4.5 mg/mL.

Preferably, the content of the ginkgo biloba extract is 2.8-4.2 mg/mL; more preferably, the content of the ginkgo biloba extract is 3.0-4.0 mg/mL; and, most preferably, the content of the ginkgo biloba extract is 3.2-3.8 mg/mL.

In a specific embodiment, the content of the ginkgo biloba extract is 2.8-4.2 mg/mL.

The injection solution of the present invention is prepared by mixing the ginkgo biloba extract with EGB 761-ginkgo biloba extract.

As EGB761 folium Ginkgo extract, the total flavonoid glycoside content is not less than 24%, and the total lactone content is not less than 6%.

Preferably, the content of total flavonoid glycoside is not less than 24.2%, and the content of total lactone is not less than 6.3%; more preferably, the content of total flavonoid glycosides is not less than 24.4% and the content of total lactones is not less than 6.6%; and, most preferably, the total flavonoid glycoside content is not less than 24.6% and the total lactone content is not less than 7.0%.

In a specific embodiment, the EGB761 ginkgo biloba extract has a total flavonoid glycoside content of 24.7% and a total lactone content of 7.2%.

According to the injection of the invention, the proportion of the ginkgolide B in the total lactones is not less than 5%; the proportion of bilobalide in total lactone is not less than 40%.

Preferably, the proportion of the ginkgolide B in the total lactones is not less than 8%; the proportion of bilobalide in the total lactone is not less than 42%; more preferably, the proportion of ginkgolide B in total lactones is not less than 12%; the proportion of bilobalide in the total lactone is not less than 45%; and, most preferably, the proportion of ginkgolides B in total lactones is not less than 15%; the proportion of bilobalide in total lactone is not less than 50%.

In a specific embodiment, the proportion of ginkgolides B to total lactones is 17.6% and the proportion of bilobalide to total lactones is 51.3%.

The injection solution according to the present invention, wherein the injection solution further comprises alkyl glucoside.

Preferably, the alkyl glucoside is selected from C6-C14 alkyl glucoside; more preferably, the alkyl glucoside is selected from the group consisting of C6-C12 alkyl glucosides; and, most preferably, the alkyl glucoside is selected from the group consisting of C8-C12 alkyl glucosides.

In a specific embodiment, the alkyl glucoside is selected from decyl glucoside.

The injection solution of the present invention, wherein the average degree of polymerization of the alkyl glucoside is 1.1 to 2.2.

Preferably, the average degree of polymerization of the alkyl glucoside is 1.2-2.0; more preferably, the average degree of polymerization of the alkyl glucoside is 1.3 to 1.9; and, most preferably, the average degree of polymerization of the alkyl glucoside is preferably 1.4 to 1.8.

In a specific embodiment, the alkyl glucoside has an average degree of polymerization of 1.6.

According to the injection of the present invention, the weight ratio of the ginkgo biloba extract to the alkyl glucoside is 1: (6-10).

Preferably, the weight ratio of the ginkgo biloba extract to the alkyl glucoside is 1: (6.4-9.6); more preferably, the weight ratio of the ginkgo biloba extract to the alkyl glucoside is 1: (6.8-9.2); and, most preferably, the weight ratio of the ginkgo biloba extract to the alkyl glucoside is 1: (7.2-8.8).

In a specific embodiment, the weight ratio of the ginkgo biloba extract to the alkyl glucoside is 1: 8.

The injection solution according to the present invention, wherein the injection solution further comprises ethyl oleate and oleic acid.

Preferably, the weight ratio of the ginkgo biloba extract to the ethyl oleate and the oleic acid is 1: (15-35): (0.5-2.5); more preferably, the weight ratio of the ginkgo biloba extract to the ethyl oleate and the oleic acid is 1: (20-32): (0.8-2.0); and, most preferably, the weight ratio of the ginkgo biloba extract to the ethyl oleate and the oleic acid is 1: (23-29): (1.0-1.7).

In a specific embodiment, the weight ratio of the ginkgo biloba extract to the ethyl oleate and the oleic acid is 1: 22.8: 1.1.

the injection solution according to the present invention comprises the following components:

2.8-4.2g of ginkgo leaf extract;

60-110g of ethyl oleate;

26.9-28g of alkyl glucoside;

3-7g of oleic acid;

40-60mL of absolute ethyl alcohol;

20-40mL of 1, 3-propylene glycol;

water for injection to 1000 mL.

In a specific embodiment, the composition of the injection solution is as follows:

2.8-4.2g of EGB761 ginkgo leaf extract;

60-110g of ethyl oleate;

decyl glucoside (average degree of polymerization of 1.4-1.8) 26.9-28 g;

3-7g of oleic acid;

40-60mL of absolute ethyl alcohol;

20-40mL of 1, 3-propylene glycol;

water for injection to 1000 mL.

In another aspect, the present invention also provides a preparation method of the aforementioned ginkgo biloba extract injection according to the present invention, which comprises:

adding folium Ginkgo extract and ethyl oleate into anhydrous ethanol, heating to 50-60 deg.C, stirring to mix well, adding decyl glucoside, and mixing well to obtain oil phase;

adding 1, 3-propylene glycol and oleic acid in a formula amount into 400-600mL of injection water, heating to 50-60 ℃, and uniformly stirring to obtain a water phase;

adding the oil phase into the water phase under the high-speed shearing action of 20000rpm, and continuously shearing at high temperature for 1-10min to obtain primary emulsion;

adding the rest of water for injection into the colostrum, and adjusting the pH value to 5.5-6.5; then transferring the ginkgo biloba extract micro-emulsion to a high-pressure homogenizing machine, controlling the pressure at 800-1200MPa, the temperature at 40-50 ℃, and homogenizing for 4-8 times to obtain the ginkgo biloba extract micro-emulsion;

filtering and sterilizing with 0.22 μm microporous membrane, and packaging into penicillin bottles at a ratio of 5 mL/piece to obtain folium Ginkgo extract injection.

Without wishing to be bound by any theory, the ginkgo biloba extract of the present invention has higher encapsulation efficiency of ginkgolide B and bilobalide, which respectively reach 85% and 90% above, due to the addition of alkyl glucoside; and meanwhile, the particle size is smaller. When the ginkgo biloba extract injection is placed for a long time, decyl glucoside is not added or the addition amount is less, the chemical stability of the ginkgolide B and the bilobalide is poor, the particle size is remarkably increased, and the chemical stability of the ginkgo biloba extract injection of the embodiments 1 to 3 of the invention is remarkably increased.

Compared with the prior art, the invention has the following beneficial technical effects:

i) the ginkgo biloba extract injection has high encapsulation rate of ginkgolide B and bilobalide, has the content larger than that of the injection on the market, and simultaneously has excellent dispersion stability and chemical stability.

ii) the preparation method is simple and easy to implement, has good repeatability, and does not need other auxiliary materials; the equipment cost is low and no pollution is caused; can generate huge social benefit and economic benefit, and is suitable for being widely popularized and used.

Detailed Description

The invention will be further illustrated with reference to specific embodiments. It should be understood that these examples are for illustrative purposes only and are not intended to limit the scope of the present invention. Further, it should be understood that various changes or modifications can be made by those skilled in the art after reading the contents of the present invention, and those equivalents also fall within the scope of the invention defined by the appended claims.

The following examples will aid understanding of the present invention, but are not intended to limit the scope of the present invention.

In the present invention, the EGB761 ginkgo biloba extract is obtained from Zhejiang Connbei pharmaceutical Co. The content of total flavonoid glycoside in the folium Ginkgo extract is 24.7%, and the content of total lactone is 7.2%; wherein, the proportion of ginkgolide B in total lactone is 17.6%, and the proportion of bilobalide in total lactone is 51.3%.

Example 1:

3.5g of EGB761 ginkgo leaf extract;

100g of ethyl oleate;

decyl glucoside (average degree of polymerization of 1.6) 28 g;

4g of oleic acid;

50mL of absolute ethyl alcohol;

30mL of 1, 3-propylene glycol;

water for injection is 1000 mL;

adding EGB761 folium Ginkgo extract and ethyl oleate into anhydrous ethanol, heating to 60 deg.C, stirring to mix well, adding decyl glucoside, and mixing well to obtain oil phase;

adding 1, 3-propylene glycol and oleic acid in a formula amount into 500mL of water for injection, heating to 60 ℃, and uniformly stirring to obtain a water phase;

adding the oil phase into the water phase under the high-speed shearing action of 20000rpm, and continuously shearing at high temperature for 5min to obtain primary emulsion;

adding the rest of water for injection into the colostrum, and adjusting the pH value to 6.0; then transferring the ginkgo biloba extract micro-emulsion into a high-pressure homogenizing machine, controlling the pressure to be 800MPa and the temperature to be 50 ℃, and homogenizing for 6 times to obtain ginkgo biloba extract micro-emulsion;

the extract injection of ginkgo biloba leaves of example 1 was obtained by filtration sterilization using a 0.22 μm microfiltration membrane and dispensing 5 mL/vial.

Example 2:

2.8g of EGB761 ginkgo leaf extract;

64g of ethyl oleate;

decyl glucoside (average degree of polymerization of 1.4) 26.9 g;

3g of oleic acid;

40mL of absolute ethyl alcohol

1, 3-propanediol 20mL

Water for injection to 1000mL

Adding EGB761 folium Ginkgo extract and ethyl oleate into anhydrous ethanol, heating to 50 deg.C, stirring to mix well, adding decyl glucoside, and mixing well to obtain oil phase;

adding 1, 3-propylene glycol and oleic acid in a formula amount into 400mL of water for injection, heating to 50 ℃, and uniformly stirring to obtain a water phase;

adding the oil phase into the water phase under the high-speed shearing action of 20000rpm, and continuously shearing at high temperature for 5min to obtain primary emulsion;

adding the rest of water for injection into the colostrum, and adjusting the pH value to 6.5; then transferring the ginkgo biloba extract micro-emulsion into a high-pressure homogenizing machine, controlling the pressure to be 1000MPa and the temperature to be 40 ℃, and homogenizing for 8 times to obtain ginkgo biloba extract micro-emulsion;

the extract injection of ginkgo biloba leaves of example 1 was obtained by filtration sterilization using a 0.22 μm microfiltration membrane and dispensing 5 mL/vial.

Example 3:

4.2g of EGB761 ginkgo leaf extract;

110g of ethyl oleate;

decyl glucoside (average degree of polymerization of 1.8) 26.9 g;

oleic acid 7 g;

60mL of absolute ethyl alcohol;

40mL of 1, 3-propylene glycol;

water for injection is 1000 mL;

adding EGB761 folium Ginkgo extract and ethyl oleate into anhydrous ethanol, heating to 60 deg.C, stirring to mix well, adding decyl glucoside, and mixing well to obtain oil phase;

adding 1, 3-propylene glycol and oleic acid in a formula amount into 600mL of water for injection, heating to 60 ℃, and uniformly stirring to obtain a water phase;

adding the oil phase into the water phase under the high-speed shearing action of 20000rpm, and continuously shearing at high temperature for 5min to obtain primary emulsion;

adding the rest of water for injection into the colostrum, and adjusting the pH value to 5.5; then transferring the ginkgo biloba extract micro-emulsion into a high-pressure homogenizing machine, controlling the pressure to be 1200MPa and the temperature to be 60 ℃, and homogenizing for 4 times to obtain ginkgo biloba extract micro-emulsion;

the extract injection of ginkgo biloba leaves of example 1 was obtained by filtration sterilization using a 0.22 μm microfiltration membrane and dispensing 5 mL/vial.

Comparative example 1:

the same as example 1, but only 10g of decyl glucoside were added.

Comparative example 2:

same as example 1, but without decyl glucoside.

Comparative example 3:

the decyl glucoside was replaced with the same weight of poloxamer 188 as in example 1.

The contents of ginkgolide B and bilobalide and their average particle diameters of the ginkgo biloba extract injections of examples 1 to 3 of the present invention and comparative examples 1 to 3 were measured according to the following methods, respectively. The content test method of ginkgolide B and bilobalide adopts an HPLC method, and the conditions are as follows: agilent Eclips C₁₈A chromatographic column with specification of 4.6mm multiplied by 250mm is multiplied by 5 mu m; the detection wavelength is 360 nm; the flow rate is 1 mL/min; the mobile phase is a mixed solution of methanol, tetrahydrofuran and water according to the volume ratio of 30:10: 60; the column temperature is 30 ℃; the amount of the sample was 10. mu.L. During the test, the ginkgolide B and the bilobalide reference substance are respectively prepared into 500 mug/mL reference substance solutions by using acetone, and the ginkgo biloba extract injection of the embodiments 1 to 3 and the comparative examples 1 to 3 of the invention are diluted into the solution by using the acetone as the test solution. 10 μ L of each of the reference solution and the sample solution is taken to record chromatographic peak, and the contents of ginkgolide B and bilobalide are calculated according to the area of the chromatographic peak. In the average particle size measurement, a small amount of the ginkgo biloba extract injection of examples 1 to 3 and comparative examples 1 to 3 was added to water, uniformly dispersed by shaking, and the average particle size was measured using a laser particle sizer. The results are specifically shown in Table 1.

TABLE 1

Then, the mixture was left for 1 month at 40 ℃ and 75% relative humidity, filtered through a 0.45 μm microporous membrane, and the content of bilobalide B and bilobalide and the average particle diameter were measured for 5 days, 10 days and 30 days. The results are specifically shown in Table 2.

TABLE 2

The results show that the ginkgo biloba extract of the embodiments 1 to 3 of the invention has higher encapsulation rates of the ginkgolide B and the bilobalide due to the addition of decyl glucoside, the encapsulation rates of the ginkgolide B and the bilobalide are respectively more than 85 percent and 90 percent, and the content of the ginkgo biloba extract is more than that of the injection on the market; and meanwhile, the particle size is smaller. When the ginkgo biloba extract injection is placed for a long time, decyl glucoside is not added or the addition amount is less, the stability of the ginkgolide B and the bilobalide is poor, the particle size is remarkably increased, the chemical stability of the ginkgo biloba extract injection of the embodiments 1 to 3 of the invention is remarkably increased, and the content change is less than 4%; the average particle size also remained essentially unchanged.

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 shall fall within the protection scope of the present invention.

Claims (6)

1. The injection of the ginkgo biloba extract is microemulsion of the ginkgo biloba extract, and is characterized by comprising the following components:

2.8-4.2g of ginkgo leaf extract;

60-110g of ethyl oleate;

26.9-28g of alkyl glucoside;

3-7g of oleic acid;

40-60mL of absolute ethyl alcohol;

20-40mL of 1, 3-propylene glycol;

water for injection to 1000 mL.

2. The injection according to claim 1, wherein the ginkgo biloba extract is selected from the group consisting of EGB 761-ginkgo biloba extract.

3. The injection solution of claim 1, wherein the alkyl glucoside is selected from the group consisting of C6-C14 alkyl glucosides.

4. The injection according to claim 1, wherein the alkyl glucoside has an average degree of polymerization of 1.1-2.2.

5. The injection solution according to claim 1, wherein the composition of the injection solution is as follows:

2.8-4.2g of EGB761 ginkgo leaf extract;

60-110g of ethyl oleate;

the average polymerization degree is 1.4-1.8, and the average polymerization degree is 26.9-28g of decyl glucoside;

3-7g of oleic acid;

40-60mL of absolute ethyl alcohol;

20-40mL of 1, 3-propylene glycol;

water for injection to 1000 mL.

6. A method for preparing the ginkgo biloba leaf extract injection according to claim 5, comprising:

adding folium Ginkgo extract and ethyl oleate into anhydrous ethanol, heating to 50-60 deg.C, stirring to mix well, adding decyl glucoside, and mixing well to obtain oil phase;

adding 1, 3-propylene glycol and oleic acid in a formula amount into 400-600mL of injection water, heating to 50-60 ℃, and uniformly stirring to obtain a water phase;

adding the oil phase into the water phase under the high-speed shearing action of 20000rpm, and continuously shearing at high temperature for 1-10min to obtain primary emulsion;

adding the rest of water for injection into the colostrum, and adjusting the pH value to 5.5-6.5; then transferring the ginkgo biloba extract micro-emulsion to a high-pressure homogenizing machine, controlling the pressure at 800-1200MPa, the temperature at 40-50 ℃, and homogenizing for 4-8 times to obtain the ginkgo biloba extract micro-emulsion;

filtering and sterilizing with 0.22 μm microporous membrane, and packaging in penicillin bottles at 5 mL/branch to obtain the extract injection of folium Ginkgo according to claim 5.