Nanoemulsion containing tamoxifen and ginsenoside and preparation method thereof

Abstract

The invention relates to the technical field of pharmaceutical preparations, in particular to a nanoemulsion containing tamoxifen and ginsenoside and a preparation method of the nanoemulsion. The tamoxifen citrate, the ginsenoside, the tributyrin, the isopropyl myristate, the tragacanth gum, the poloxamer and the glycerol are prepared into the nano-emulsion, the prepared nano-emulsion has a small particle size and good stability, the solubility can be remarkably improved, and the targeting property and the treatment effect of the nano-emulsion are further improved.

Description

Technical Field

[0001] The invention belongs to the technical field of preparations, and particularly relates to a nanoemulsion containing tamoxifen and ginsenosides and a preparation method thereof. Background Art

[0002] Breast cancer is the second most common malignant tumor after lung cancer. The estrogen drug tamoxifen has been widely used in the clinical treatment of advanced breast cancer and ovarian cancer. Its anti-cancer mechanism is the inhibition of estrogen receptors and cytotoxicity, and the efficacy can be as high as 49%. However, with the widespread use of tamoxifen, breast cancer cells have shown multidrug resistance to it, which greatly limits the anti-cancer efficacy of tamoxifen.

[0003] Ginsenosides are natural anti-cancer ingredients that scientists have high hopes for. Studies have shown that ginsenosides exert anti-cancer effects through multiple mechanisms, including inhibiting tumor cell proliferation, inducing tumor cell apoptosis, inhibiting tumor cell migration and invasion, inhibiting tumor angiogenesis, enhancing the body's immune function, and resisting oxidative stress. These mechanisms work together to make ginsenosides show significant potential in cancer treatment.

[0004] However, not only is tamoxifen poorly soluble and has poor drug resistance, but ginsenosides also have a low solubility in water and are almost insoluble in water, which directly affects the absorption and utilization of the drug and greatly limits the efficacy of the drug.

[0005] Therefore, how to improve the drug resistance of tamoxifen and increase the solubility of the drug is the key to improving the efficacy.

[0006] At present, the only marketed dosage forms of tamoxifen and ginsenosides are oral preparations, and according to the dissolution test standards of the 2020 edition of the "Chinese Pharmacopoeia", the 30-minute dissolution amount of ordinary tamoxifen citrate tablets is only 75% of the labeled amount, which does not effectively solve the dissolution problem.

[0007] CN111925295A discloses a tamoxifen citrate-ginsenoside co-amorphous product. In a dissolution test, using a 0.1M hydrochloric acid medium, the release of tamoxifen in 60 minutes reached 90%, and the release of ginsenoside Rg in 60 minutes reached 88%. However, the problem of poor stability of the prepared tamoxifen citrate-ginsenoside co-amorphous product was not solved. Summary of the invention

[0008] In view of the deficiencies in the prior art, the technical problem to be solved by the present invention is to provide a tamoxifen citrate-ginsenoside nanoemulsion and a preparation method and use thereof in view of the defects of low solubility and dissolution rate of the above-mentioned drugs.

[0009] The technical solution adopted by the present invention to solve the technical problem is: preparing a composite nanoemulsion containing tamoxifen, preferably, the tamoxifen nanoemulsion also contains ginsenoside.

[0010] Furthermore, the weight proportions of the components of the tamoxifen composite nanoemulsion are:

[0011] 10-15 parts by weight of tamoxifen citrate;

[0012] 8-10 parts by weight of ginsenosides;

[0013] Tributyrin 25-50 parts by weight;

[0014] Isopropyl myristic acid 30-60 parts by weight;

[0015] 60-100 parts by weight of tragacanth gum;

[0016] Poloxamer 20-50 parts by weight;

[0017] Glycerol 60-100 parts by weight.

[0018] Furthermore, the weight proportions of the components of the tamoxifen nanoemulsion are:

[0019] 10-15 parts by weight of tamoxifen citrate;

[0020] 8-10 parts by weight of ginsenosides;

[0021] Tributyrin 25-35 parts by weight;

[0022] Isopropyl myristic acid 30-50 parts by weight;

[0023] 60-80 parts by weight of tragacanth gum;

[0024] Poloxamer 20-35 parts by weight;

[0025] Glycerol 60-100 parts by weight.

[0026] The tamoxifen nanoemulsion preparation process comprises the following steps:

[0027] (1) dissolving tamoxifen and ginsenosides in an appropriate amount of ethanol, mixing tributyrin and isopropyl myristate in a certain proportion and stirring evenly, slowly adding the dissolved drug solution into the oil phase, and sonicating to prepare an oil phase mixed solution;

[0028] (2) After mixing tragacanth gum and poloxamer evenly, add glycerol, heat and stir evenly to prepare an emulsifier mixture;

[0029] (3) slowly adding the oil phase mixture of step (1) into the emulsifier mixture of step (2), stirring evenly, and further ultrasonically refining after forming colostrum;

[0030] (4) adding an appropriate amount of water to the colostrum refined by ultrasonication in step (3), stirring evenly to form a coarse emulsion, and homogenizing to prepare a tamoxifen composite nanoemulsion.

[0031] Preferably, the ultrasonic power in step (1) is 30% to 35%.

[0032] Preferably, a surfactant is further added in step (2), specifically a polyvinyl alcohol aqueous solution with a concentration of 1% to 2%.

[0033] Preferably, in step (2), the pH value of the emulsifier mixture is adjusted to 5.0-6.5, and the heating temperature is adjusted to 40-60°C.

[0034] Preferably, when the organic phase and the aqueous phase are mixed in step (3), the volume ratio of the organic phase to the aqueous phase is 3:1 to 6:1.

[0035] Preferably, in step (3), the homogenization pressure is set to 18000-29000 psi, the homogenization is performed 2-3 times, and the temperature is maintained at 20±3°C during the homogenization process.

[0036] Specifically, the tamoxifen nanoemulsion preparation process comprises the following steps:

[0037] (1) taking appropriate amounts of tributyrin and isopropyl myristate, adding tamoxifen and ginsenoside, and ultrasonicating at a power of 30% to 35%, a temperature of 55 to 65° C., and a time of 0.5 to 1 h to prepare an oil phase mixture;

[0038] (2) After mixing tragacanth gum and poloxamer, add glycerol and a 1% to 2% aqueous solution of polyvinyl alcohol, adjust the pH value to 5.0-6.5, heat at 40-60° C., stir evenly, and prepare an emulsifier mixture;

[0039] (3) slowly adding the oil phase mixture of step (1) into the emulsifier mixture of step (2), wherein the volume ratio of the organic phase to the aqueous phase is 3:1 to 6:1, stirring evenly, and further ultrasonically refining after forming colostrum;

[0040] (4) adding an appropriate amount of water to the colostrum refined by ultrasonication in step (3), stirring evenly to form a coarse emulsion, and homogenizing the colostrum. The pressure during the homogenization process is set to 18000-29000 psi, and the homogenization is performed 2-3 times. The temperature is maintained at 20±3° C. during the homogenization process to prepare a tamoxifen composite nanoemulsion.

[0041] Furthermore, the step (4) further comprises a centrifugation step, specifically, adding an appropriate amount of water to the colostrum refined by ultrasound in step (3), stirring evenly to form a crude milk, homogenizing, discarding the supernatant, centrifuging at high speed, washing the precipitate with water, drying, and adding auxiliary materials to prepare a pharmaceutically acceptable preparation.

[0042] Preferably, the pharmaceutically acceptable preparation is one or more of nanoemulsion gel, soft capsule, tablet, capsule, injection.

[0043] Furthermore, the present invention also provides pharmaceutical uses of the tamoxifen composite nanoemulsion, specifically, uses of the tamoxifen composite nanoemulsion in preparing drugs for preventing and treating breast cancer.

[0044] The present invention has the following beneficial effects:

[0045] (1) Tamoxifen citrate and ginsenosides are prepared into the same nanoformulation, which has a simple preparation process and significantly improves the solubility and bioavailability of the two drugs;

[0046] (2) The prepared nanoemulsion has a small particle size and good stability;

[0047] (3) Pharmacological experiments have confirmed that it can significantly reduce drug irritation and side effects, and further improve the targeting and therapeutic effects of nanoemulsions. BRIEF DESCRIPTION OF THE DRAWINGS

[0048] Figure 1 Total impurity content in the stability test of tamoxifen nanoemulsion prepared by the present invention

[0049] Figure 2 Cumulative dissolution of tamoxifen in the tamoxifen nanoemulsion prepared by the present invention

[0050] Figure 3 Cumulative dissolution of ginsenosides in tamoxifen nanoemulsion prepared by the present invention DETAILED DESCRIPTION

[0051] The present invention is further described below by way of examples. It should be correctly understood that the examples of the present invention are only used to illustrate the present invention, rather than to limit the present invention, and those skilled in the art should be aware that the examples described herein do not limit the present invention in any way.

[0052] Embodiment 1:

[0053] Composition ratio: 10 parts by weight of tamoxifen citrate;

[0054] 10 parts by weight of ginsenosides;

[0055] Tributyrin 35 parts by weight;

[0056] Isopropyl myristate 50 parts by weight;

[0057] 60 parts by weight of tragacanth gum;

[0058] Poloxamer 20 parts by weight;

[0059] 60 parts by weight of glycerol;

[0060] Preparation method:

[0061] (1) taking appropriate amounts of tributyrin and isopropyl myristate, adding tamoxifen and ginsenoside, and ultrasonicating at a power of 30% to 35%, a temperature of 60° C., and a time of 0.5 h to prepare an oil phase mixture;

[0062] (2) After mixing tragacanth gum and poloxamer, add glycerol and a 1% polyvinyl alcohol aqueous solution, adjust the pH value to 5.5, heat at 50° C., and stir evenly to prepare an emulsifier mixture;

[0063] (3) slowly adding the oil phase mixture of step (1) into the emulsifier mixture of step (2), wherein the volume ratio of the organic phase to the aqueous phase is 5:1, stirring evenly, and further ultrasonically refining after forming colostrum;

[0064] (4) adding an appropriate amount of water to the colostrum refined by ultrasonication in step (3), stirring evenly to form a coarse emulsion, and homogenizing the colostrum. The pressure during the homogenization process is set to 18000-29000 psi, and the homogenization is performed twice. The temperature is maintained at 202° C. during the homogenization process to prepare a tamoxifen composite nanoemulsion.

[0065] Embodiment 2:

[0066] Composition ratio: Tamoxifen citrate 15g;

[0067] Ginsenoside 8g;

[0068] Tributyrin 30g;

[0069] Isopropyl myristate 50g;

[0070] Tragacanth gum 60g;

[0071] Poloxamer 35g;

[0072] Glycerol 100g;

[0073] Preparation method:

[0074] (1) taking appropriate amounts of tributyrin and isopropyl myristate, adding tamoxifen and ginsenoside, and ultrasonicating at a power of 30% to 35%, a temperature of 55° C., and a time of 1 h to prepare an oil phase mixture;

[0075] (2) After mixing tragacanth gum and poloxamer, add glycerol and a 2% polyvinyl alcohol aqueous solution, adjust the pH value to 6.5, heat at 60° C., and stir evenly to prepare an emulsifier mixture;

[0076] (3) slowly adding the oil phase mixture of step (1) into the emulsifier mixture of step (2), wherein the volume ratio of the organic phase to the aqueous phase is 6:1, stirring evenly, and further ultrasonically refining after forming colostrum;

[0077] (4) adding an appropriate amount of water to the colostrum refined by ultrasonication in step (3), stirring evenly to form a coarse emulsion, and homogenizing the colostrum. The pressure during the homogenization process is set to 29000 psi, and the homogenization is performed twice. The temperature is maintained at 202° C. during the homogenization process to prepare a tamoxifen composite nanoemulsion.

[0078] Example 3

[0079] Components of each group: 15 g of tamoxifen citrate;

[0080] Ginsenoside 10g;

[0081] Tributyrin 35g;

[0082] Isopropyl myristate 30g;

[0083] Tragacanth gum 80g;

[0084] Poloxamer 35g;

[0085] Glycerol 80g

[0086] The preparation method is the same as Example 2.

[0087] Example 4

[0088] Components of each group: 15 g of tamoxifen citrate;

[0089] Ginsenoside 10g;

[0090] Tributyrin 25g;

[0091] Isopropyl myristate 60g;

[0092] Tragacanth gum 80g;

[0093] Poloxamer 50g;

[0094] Glycerol 100g

[0095] The preparation method is the same as Example 1.

[0096] Example 5

[0097] Components of each group: 30 g of tamoxifen citrate;

[0098] Ginsenoside 16g;

[0099] Tributyrin 50g;

[0100] Isopropyl myristate 60g;

[0101] Tragacanth gum 100g;

[0102] Poloxamer 20g;

[0103] Glycerol 60g

[0104] The preparation method is the same as Example 2.

[0105] Example 6

[0106] Take appropriate amounts of tamoxifen citrate and ginsenosides, and other components according to the weight portions in the following table. The specific operating steps are carried out according to the method in Example 1.

[0107] Table 1 Tamoxifen nanoemulsions with different ratios

[0108]

[0109] Table 2 Tamoxifen nanoemulsions with different components

[0110]

[0111]

[0112] The prepared nanoemulsion was tested for stability and particle size. The accelerated stability test was conducted at 40°C ± 2°C and 75% ± 5% relative humidity for 6 months, and the precipitation, stratification, particle size and impurities were recorded.

[0113] Table 3 Stability test results

[0114]

[0115] From the results in the table above, it can be seen that nanoemulsion 4 and nanoemulsion 8 showed excellent stability, while other groups showed precipitation, stratification or large changes in particle size to varying degrees. At the same time, the impurity situation is shown in Table 1. It can be seen that after 6 months of experiments, the total impurities in each group increased to varying degrees, among which nanoemulsion 4 and nanoemulsion 8 had the lowest total impurities.

[0116] Example 7 Dissolution Test

[0117] Nanoemulsions 6-9 in Examples 1-3 and 6 were taken, centrifuged at high speed, precipitated, washed with water, dried, and added with auxiliary materials to prepare tablets. The dissolution was carried out according to the quality standard of tamoxifen citrate tablets in the 2020 edition of the Chinese Pharmacopoeia.

[0118] Tablets prepared from nanoemulsions 6-9 in Examples 1-3 and 6 were taken separately, and the dissolution medium was 900 mL of 0.1 M hydrochloric acid, the rotation speed was 75 r / min, and the temperature was (37±0.5)°C, and the solubility of ginsenoside Re therein was calculated.

[0119] Specifically, the dissolution rate of tamoxifen is Figure 2 As shown in Figure 2, the solubility of ginsenosides is Figure 3 As shown. Figure 2 The dissolution rate of tamoxifen in Example 1-3 was higher than 88% at 30 min, and almost all of it was dissolved at 60 min. Figure 2 The solubility of ginsenoside Re in Examples 1-3 is higher than 82% at 30 min and higher than 92% at 60 min.

[0120] Example 8 Effect of the composition of the present invention on breast cancer mouse model

[0121] 1 Experimental Materials

[0122] 1.1 Experimental animals

[0123] Healthy BABL / c mice (provided by the Pharmacology Center of Lunan Pharmaceutical Group) weighed 20-22 g and began the experiment after 7 days of adaptive feeding.

[0124] 1.2 Experimental Reagents

[0125] Commercially available tamoxifen citrate tablets, Example 3 tamoxifen nanoemulsion.

[0126] 2 Test methods

[0127] 2.1 Modeling

[0128] Healthy BABL / c mice were subcutaneously inoculated with 4T1 breast cancer cell suspension in the fourth mammary fat pad on the left side of the mice to establish a mouse breast cancer model. The concentration was 2*10 6 / mL, each mouse was inoculated with 0.1mL for 7 consecutive days.

[0129] 2.2 Grouping

[0130] The successfully modeled mice were divided into a model group, an Example 3 group, and a tamoxifen citrate tablet group, with 12 mice in each group. The drugs were administered when tumor nodules appeared at the site of 4T1 breast cancer cell inoculation. The tamoxifen group in Example 3 was administered with the nanoemulsion of Example 3 (4 mg / kg) by gavage, and the tamoxifen citrate tablet group was administered with tamoxifen citrate tablets (6 mg / kg) by gavage. The model group was administered with an equal volume of normal saline, and the drugs were administered continuously for 30 days.

[0131] 2.3 Measurement of tumor volume and mouse body weight

[0132] 2.4 Data Processing

[0133] SPSS 22.0 software was used for statistical analysis, and the data were expressed as “mean ± standard deviation”. One-way analysis of variance was used for analysis among multiple groups, and P < 0.05 was considered statistically significant.

[0134] 3 Test results

[0135] 3.1 Daily observation of mice

[0136] After 30 days of daily observation, it was found that as the tumor volume of the mice in the model group increased, the mice showed reduced activity and decreased appetite. The mice in the Example 3 group and the commercially available tamoxifen citrate group showed a brief decrease in activity at the beginning of administration, but returned to normal after 1-2 days, with a good diet and mental state.

[0137] 3.2 Observation of mouse body weight and tumor volume

[0138] After 30 days of drug administration in each group, it can be seen from Table 4 that the weight loss trend of mice in Example 3 group was significantly slower than that in the model group and the commercially available tamoxifen citrate group. After drug administration, the tumor volume of mice in Example 3 was significantly smaller than that in other groups.

[0139] Table 4 Observation results of tumor volume of mice by weight

[0140]

[0141] The above experiments confirm that the tamoxifen nanoemulsion prepared by the present invention has a significant inhibitory effect on tumor growth in a breast cancer mouse model, can slow down the growth of tumor volume, and can significantly improve the survival status of mice.

Claims

1. A composite nanoemulsion containing tamoxifen and a preparation method thereof, characterized in that: The tamoxifen nanoemulsion also contains ginsenosides. Further, the weight parts of the components are: 10-15 parts by weight of tamoxifen citrate; 8-10 parts by weight of ginsenosides; Tributyrin 25-50 parts by weight; Isopropyl myristic acid 30-60 parts by weight; 60-100 parts by weight of tragacanth gum; Poloxamer 20-50 parts by weight; Glycerol 60-100 parts by weight.

2. The composite nanoemulsion of tamoxifen according to claim 1, characterized in that: The weight parts of the components of the tamoxifen nanoemulsion are as follows: 10-15 parts by weight of tamoxifen citrate; 8-10 parts by weight of ginsenosides; Tributyrin 25-35 parts by weight; Isopropyl myristic acid 30-50 parts by weight; 60-80 parts by weight of tragacanth gum; Poloxamer 20-35 parts by weight; Glycerol 60-100 parts by weight.

3. The tamoxifen composite nanoemulsion according to claim 1, characterized in that: The following steps are involved: (1) taking appropriate amounts of tributyrin and isopropyl myristate, adding tamoxifen and ginsenoside, and sonicating to prepare an oil phase mixture; (2) After mixing tragacanth gum and poloxamer evenly, add glycerol, heat and stir evenly to prepare an emulsifier mixture; (3) slowly adding the drug solution of step (2) into the oil phase mixture of step (1), stirring evenly, and further ultrasonically refining after forming colostrum; (4) adding an appropriate amount of water to the colostrum refined by ultrasonication in step (3), stirring evenly to form a coarse emulsion, and homogenizing to prepare a tamoxifen composite nanoemulsion.

4. The tamoxifen composite nanoemulsion according to claim 3, characterized in that: The ultrasonic power in step (2) is 15% to 30%.

5. The tamoxifen composite nanoemulsion according to claim 3, characterized in that: In the step (2), the pH value of the emulsifier mixture is adjusted to 5.0-6.5, and the heating temperature is increased to 40-60°C.

6. The tamoxifen composite nanoemulsion according to claim 3, characterized in that: When the organic phase and the aqueous phase are mixed in the step (3), the volume ratio of the organic phase to the aqueous phase is 3:1 to 6:

1.

7. The tamoxifen composite nanoemulsion according to claim 3, characterized in that: In the step (3), the homogenization pressure is set to 18000-29000 psi, and the homogenization is performed 2-3 times. The temperature is maintained at 20±3°C during the homogenization process.

8. The tamoxifen composite nanoemulsion according to claim 3, characterized in that: The step (4) also includes a centrifugation step, specifically: adding an appropriate amount of water to the colostrum refined by ultrasound in step (3), stirring evenly to form a crude milk, homogenizing, high-speed centrifuging, discarding the supernatant, washing the precipitate with water, drying, and adding auxiliary materials to prepare a pharmaceutically acceptable preparation.

9. The tamoxifen composite nanoemulsion according to claim 8, characterized in that: The preparation is one or more of nanoemulsion gel, soft capsule, tablet, capsule and injection.

10. Use of the tamoxifen composite nanoemulsion according to claim 1 in preparing drugs for preventing and treating breast cancer.

Images