CN115252551A - Microemulsion and freeze-dried powder for paclitaxel injection and preparation method thereof - Google Patents

Abstract

The invention belongs to the technical field of pharmaceutical preparations, and particularly relates to microemulsion and freeze-dried powder for paclitaxel injection and a preparation method thereof. The microemulsion for paclitaxel injection comprises the following components in parts by weight: 4-8 parts of paclitaxel, 235-300 parts of oil phase, 210-275 parts of surfactant, 70-90 parts of cosurfactant and 1900-2500 parts of solvent for injection, the product does not contain polyoxyethylene castor oil and has low toxicity, and the dosage of the surfactant and the cosurfactant is small, so that the toxic and side effects of chemotherapy on patients can be greatly reduced, and the anticancer effect and the cancer cure rate are enhanced; the microemulsion for paclitaxel injection has the particle size of 40-80 nm and good stability in blood plasma. The invention adopts the water drop method to prepare the microemulsion for paclitaxel injection, has simple and convenient production process, is suitable for large-scale production of pharmaceutical enterprises, and can greatly reduce the production cost and the medical expense of patients.

Description

A kind of paclitaxel injection microemulsion, freeze-dried powder and preparation method thereof

technical field

The invention belongs to the technical field of pharmaceutical preparations, and in particular relates to paclitaxel microemulsion for injection, freeze-dried powder and a preparation method thereof.

Background technique

Paclitaxel (PTX for short) is a natural alkaloid with a molecular formula of C ₄₇ H ₅₁ NO ₁₄ and a molecular weight of 853.91. Paclitaxel, as a broad-spectrum anticancer agent, has a broad therapeutic spectrum and is indicated for ovarian cancer, breast cancer, non-small cell lung cancer, gastric cancer, head and neck cancer, Kaposi's sarcoma, and advanced forms of acute leukemia. Paclitaxel has a unique anti-tumor mechanism. It binds to tubulin in cells and promotes its polymerization, hindering tubulin dissociation, thereby arresting cell mitosis in G2/M phase and promoting apoptosis of tumor cells. However, the water solubility of paclitaxel is extremely small (<0.1 μg/mL), which makes administration difficult, and oral absorption is almost ineffective. Currently, paclitaxel can only be effective through injection.

由Cremophor EL(聚氧乙烯蓖麻油)和无水乙醇50:50(v/v)组成，用药之前需要进行脱敏处理，易造成肾毒性和心脏毒性等，毒副作用非常大。中国专利CN101612121A公开了一种紫杉醇微乳的制备方法，以精制玉米油、大豆油、橄榄油中的一种或其中两种的混合物为油相，卵磷脂、泊洛沙姆中的一种或二者的混合物为乳化剂，无水乙醇、丙二醇中的一种或二者的混合物为助乳化剂，水为水相，制备成紫杉醇或者其衍生物的自乳化微乳，该制剂可用于口服给药，但所制备的产品不稳定，在通过胃肠道时药物容易被析出，而最终影响药物的吸收。At present, paclitaxel injection, which is domestically marketed

Composed of Cremophor EL (polyoxyethylene castor oil) and absolute ethanol 50:50 (v/v), desensitization treatment is required before medication, which is likely to cause nephrotoxicity and cardiotoxicity, etc., with very serious side effects. Chinese patent CN101612121A discloses a preparation method of paclitaxel microemulsion, using refined corn oil, soybean oil, olive oil or a mixture of two of them as the oil phase, and one or more of lecithin and poloxamer The mixture of the two is an emulsifier, one of absolute ethanol and propylene glycol or a mixture of the two is a co-emulsifier, water is the water phase, and it is prepared into a self-emulsifying microemulsion of paclitaxel or its derivatives, which can be used for oral administration However, the prepared product is unstable, and the drug is easily separated out when passing through the gastrointestinal tract, which ultimately affects the absorption of the drug.

Contents of the invention

The object of the present invention is to provide a microemulsion for paclitaxel injection and its preparation method and application in view of the existing problems. The paclitaxel microemulsion for injection of the present invention has good stability, does not precipitate after dilution, does not precipitate when passing through the gastrointestinal tract, has low toxicity, and is suitable for industrial production.

In order to achieve the above-mentioned purpose of the invention, the present invention provides the following technical solutions:

The invention provides a paclitaxel microemulsion for injection, which comprises the following components in parts by weight: 4-8 parts of paclitaxel, 235-300 parts of oil phase, 210-275 parts of surfactant, 70-90 parts of co-surfactant, 1900-2500 parts of solvent for injection.

Preferably, the oil phase includes one or more of castor oil, oleic acid, ethyl oleate, medium-chain fatty acid glycerides, isopropyl myristate and vitamin E acetate.

Preferably, the surfactant includes one or more of Tween 80, polyethylene glycol stearate, poloxamer 188 and soybean lecithin.

Preferably, the co-surfactant includes one or more of absolute ethanol, polyethylene glycol 400 and diethylene glycol monoethyl ether.

Preferably, the solvent for injection includes one or more of water for injection, physiological saline for injection and glucose solution for injection.

Preferably, the particle size of the paclitaxel microemulsion for injection is 40-80 nm.

Preferably, the pH value of the paclitaxel microemulsion for injection is 4-6.

The present invention also provides the preparation method of the paclitaxel injection microemulsion described in the above technical scheme, comprising the following steps:

The paclitaxel, some surfactants and co-surfactants are first mixed to obtain a mixed solution;

The mixed solution, the oil phase, the remaining part of the surfactant and the solvent for injection are mixed for the second time, and then filtered to obtain the microemulsion of paclitaxel for injection.

Preferably, the first mixing time is 5-15 minutes.

The present invention also provides a paclitaxel injection microemulsion freeze-dried powder obtained by drying the paclitaxel injection microemulsion.

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

The invention provides a paclitaxel microemulsion for injection, which comprises the following components in parts by weight: 4-8 parts of paclitaxel, 235-300 parts of oil phase, 210-275 parts of surfactant, 70-90 parts of co-surfactant, 1900-2500 parts of solvent for injection. The paclitaxel microemulsion for injection of the present invention forms a homogeneous thermodynamically stable system, does not precipitate medicine after being diluted with water for injection, and can also maintain stability in blood.

At the same time, the amount of surfactant and co-surfactant of the present invention is small, and the paclitaxel injection microemulsion prepared can greatly reduce the toxic and side effects of cancer chemotherapy on patients, enhance its anticancer efficacy, and enhance the cure rate of cancer.

Further, the surfactant of the present invention includes one or more of Tween 80, polyethylene glycol stearate, poloxamer 188 and soybean lecithin, and the co-surfactant includes dehydrated alcohol, polyethylene glycol One or more in ethylene glycol 400 and diethylene glycol monoethyl ether, the present invention has selected less toxic surfactant and co-surfactant, does not contain polyoxyethylene castor oil in this microemulsion, the present invention The surfactants and co-surfactants are less toxic, can greatly reduce clinical side effects and improve drug efficacy.

The particle size of the paclitaxel microemulsion for injection of the present invention is 40-80nm. Microemulsion is a thermodynamically stable dispersion system composed of oil phase, water phase, surfactant and co-surfactant in a certain proportion. The whole system is a transparent or translucent oil-water mixed system, which can promote drug absorption and enhance drug efficacy. It has good application prospects in the prevention and treatment of tumors, inhibiting the efflux of p-glycoprotein, reducing the occurrence of multidrug resistance and targeted drug delivery.

The present invention also provides the preparation method of paclitaxel microemulsion for injection described in the above technical scheme, the preparation method of the present invention is simple, is suitable for the industrialized production of pharmaceutical enterprises, and can greatly reduce the production cost of the product, and greatly reduce the cost of the patient. Medical expenses, the realization of the present invention will have huge market prospect and social value.

Description of drawings

In order to more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the following will briefly introduce the accompanying drawings required in the embodiments. Obviously, the accompanying drawings in the following description are only some of the present invention. Embodiments, for those of ordinary skill in the art, other drawings can also be obtained based on these drawings without any creative effort.

Fig. 1 is the particle size distribution figure of the paclitaxel injection microemulsion (PTX-ME) of embodiment 1 and the blank microemulsion (Blank-ME) of comparative example 1, wherein A is the blank microemulsion particle size distribution figure, and B is paclitaxel Particle size distribution diagram of microemulsion for injection (PTX-ME), C is the Zeta potential diagram of blank microemulsion and paclitaxel microemulsion for injection (PTX-ME);

Fig. 2 is the transmission electron microscope scanning figure of the paclitaxel injection microemulsion of embodiment 1;

和游离紫杉醇(Free-PTX)的体外释放结果图；Fig. 3 is the paclitaxel injection microemulsion of embodiment 1, paclitaxel injection

and the in vitro release results of free paclitaxel (Free-PTX);

Fig. 4 is the in vitro anti-cell proliferation experiment figure in Test Example 2, wherein A is the cell proliferation result after MCF-7 breast cancer cell administration 24h, B is the cell proliferation result after MCF-7 breast cancer cell administration 48h, C is the result of cell proliferation of MDA-MB-231 breast cancer cells after administration for 24 hours, and D is the result of cell proliferation of MDA-MB-231 breast cancer cells after administration of 48 hours;

紫杉醇注射用微乳(PTX-ME)给药后肿瘤体积变化图，B为实验最后各给药组肿瘤组织图；Fig. 5 is the tumor growth inhibition experiment figure in test example 2, wherein A is normal saline, commercially available preparation

Tumor volume changes after paclitaxel injection microemulsion (PTX-ME) administration, B is the tumor tissue diagram of each administration group at the end of the experiment;

的荷瘤裸鼠组织分布实验图。Fig. 6 is paclitaxel injection microemulsion and commercially available preparation in Test Example 2

Tissue distribution experiment map of tumor-bearing nude mice.

Detailed ways

The invention provides a paclitaxel microemulsion for injection, which comprises the following components in parts by weight: 4-8 parts of paclitaxel, 235-300 parts of oil phase, 210-275 parts of surfactant, 70-90 parts of co-surfactant, 1900-2500 parts of solvent for injection.

In the present invention, unless otherwise specified, the raw materials used are all commercially available products in this field.

In the present invention, the microemulsion of paclitaxel for injection preferably includes 4-5 parts by weight of paclitaxel.

In the present invention, the paclitaxel microemulsion for injection preferably includes 250-285 parts by weight of the oil phase. In the present invention, the function of the oil phase is to increase the solubility of the drug (paclitaxel), make the drug molecules exist in the oil droplets, increase the stability of the drug, and form an O/W type homogeneous thermodynamically stable system.

In the present invention, the oil phase preferably includes one or more of castor oil, oleic acid, ethyl oleate, medium-chain fatty acid glycerides (Labrafac lipophile WL1349), isopropyl myristate and vitamin E acetate species, more preferably including ethyl oleate and vitamin E acetate, the mass ratio of ethyl oleate and vitamin E acetate is preferably 1.5˜1.6:1.

In the present invention, the paclitaxel microemulsion for injection preferably includes 230-270 parts by weight of surfactant. In the present invention, the function of the surfactant is to increase the solubility of the drug and reduce the interfacial tension on the surface of the emulsion droplet.

In the present invention, the surfactant preferably includes one or more of Tween 80 (Tween80), polyethylene glycol stearate, poloxamer 188 (P188) and soybean lecithin, more preferably Including polyethylene glycol stearate, soybean lecithin and poloxamer 188, the mass ratio of polyethylene glycol stearate, soybean lecithin and poloxamer 188 is preferably 3～3.5:1.4～1.5 :1. In a specific embodiment of the present invention, the polyethylene glycol stearate is preferably polyethylene glycol-15 hydroxystearate (Solutol HS15).

In the present invention, the paclitaxel microemulsion for injection preferably includes 80-90 parts by weight of a co-surfactant. In the present invention, the function of the co-surfactant is to increase the solubility of the drug and form a stable microemulsion.

In the present invention, the co-surfactant preferably includes one or more of absolute ethanol, polyethylene glycol 400 (PEG400) and diethylene glycol monoethyl ether (Trancutol HP), more preferably diethylene glycol Alcohol monoethyl ether (Trancutol HP).

In the present invention, the paclitaxel microemulsion for injection preferably includes 1900-2000 parts by weight of solvent for injection.

In the present invention, the solvent for injection preferably includes one or more of water for injection, physiological saline for injection and glucose solution for injection, more preferably water for injection.

In the present invention, the particle size of the paclitaxel microemulsion for injection is preferably 40-80 nm, more preferably 50 nm; the shape is preferably spherical or quasi-spherical, with uniform size and uniform distribution.

In the present invention, the paclitaxel microemulsion for injection is preferably a clear, transparent O/W microemulsion with light blue opalescence.

In the present invention, the pH value of the paclitaxel microemulsion for injection is preferably 4-6. In the present invention, the pH value is preferably achieved by controlling the type and amount of each raw material, without adding a pH regulator for adjustment.

The present invention also provides the preparation method of the paclitaxel injection microemulsion described in the above technical scheme, comprising the following steps:

The paclitaxel, some surfactants and co-surfactants are first mixed to obtain a mixed solution;

The mixed solution, the oil phase, the remaining part of the surfactant and the solvent for injection are mixed for the second time, and then filtered to obtain the microemulsion of paclitaxel for injection.

In the present invention, paclitaxel, some surfactants and co-surfactants are first mixed to obtain a mixed solution.

In the present invention, the first mixing method is preferably vortex ultrasound, and the first mixing time is preferably 5-15 minutes, more preferably 10 minutes.

After the mixed solution is obtained, the present invention performs second mixing of the mixed solution, the oil phase, the remaining surfactant and the solvent for injection, and then filters to obtain the microemulsion of paclitaxel for injection.

In the present invention, it is preferred that the oil phase, the rest of the surfactant and the solvent for injection are sequentially added to the mixed solution.

In a specific embodiment of the present invention, it is preferable to add the oil phase to the mixed solution first, and the oil phase is preferably added to the mixed solution with stirring, and the present invention has no special restrictions on the time of stirring It is required that the oil phase is completely dissolved and mixed uniformly.

In the present invention, the temperature accompanied by stirring is preferably 33-40°C, more preferably 37°C.

After adding the oil phase, the present invention preferably dissolves the remaining part of the surfactant in the solvent for injection to obtain a solution for injection and then adds it to the solution. The method of adding the solution for injection is preferably dripping, that is, the water drop method .

After the oil phase and the injection solution are sequentially added to the mixed solution, a clear and transparent paclitaxel microemulsion is preferably obtained.

In the present invention, the filtration is preferably sterilized through a 0.22 μm microporous membrane.

The present invention also provides a paclitaxel injection microemulsion freeze-dried powder, which is obtained by drying the paclitaxel injection microemulsion described in the above technical scheme.

In the present invention, the drying preferably includes freeze-drying or spray-drying. The present invention has no special requirements on the conditions of the freeze-drying or spray-drying, and methods well known to those skilled in the art can be used.

In the present invention, the paclitaxel microemulsion for injection is preferably further prepared into a large infusion solution for use.

In order to further illustrate the present invention, the paclitaxel injection microemulsion, freeze-dried powder and preparation method thereof provided by the present invention are described in detail below in conjunction with the accompanying drawings and examples, but they cannot be interpreted as limiting the protection scope of the present invention.

Example 1

Prepare paclitaxel microemulsion for injection according to the prescription composition in Table 1, and make 30 bags of 100mL infusion in total:

The prescription composition of the paclitaxel injection microemulsion of table 1 embodiment 1

Paclitaxel 5.0g / ethyl oleate 175g oil phase Vitamin E Acetate 110g oil phase Polyethylene glycol stearate (SolutolHS15) 150g Surfactant Soy Lecithin 70g Surfactant Diethylene glycol monoethyl ether (TrancutolHP) 88g co-surfactant Poloxamer 188 (P188) 47g Surfactant Water for Injection 2400mL /

Preparation Process:

Weigh the prescribed amount of soybean lecithin, Trancutol HP, Solutol HS15, add the prescribed amount of paclitaxel, vortex and ultrasonically mix to obtain a mixed solution; then add the prescribed amount of oil phase, stir at a constant temperature of 37°C to make the soybean lecithin and paclitaxel completely Dissolve; Dissolve the prescribed amount of P188 in water for injection to make an aqueous solution, and drop it into the above solution under constant temperature stirring at 37°C until a clear, transparent microemulsion with blue light is formed, and pass through a 0.22 μm microporous membrane , Sterilized and depyrogenated and repacked.

The drug loading amount of paclitaxel microemulsion for injection (PTX-ME) obtained in Example 1 was 1.6 mg/g.

Example 2

The difference between this example and Example 1 is that the quality of paclitaxel is 8.0 g, and the other components and preparation process are the same as in Example 1, and the drug loading of the obtained paclitaxel injection microemulsion is 2.6 mg/g.

Example 3

The difference between this example and Example 1 is that the quality of paclitaxel is 4.0 g, and the other components and preparation process are the same as in Example 1, and the drug loading of the obtained paclitaxel injection microemulsion is 1.3 mg/g.

Comparative example 1

The difference between this comparative example and the above-mentioned Example 1 is that no paclitaxel was added to obtain a blank microemulsion (Blank-ME).

test case 1

Take the blank microemulsion (Blank-ME) of Comparative Example 1 and the paclitaxel microemulsion for injection (PTX-ME) of Example 1 respectively 200 μ L and join in a 10 mL volumetric flask, dilute to the mark with distilled water, shake gently, Particle size, polydispersity index (PDI) and Zeta potential were measured by ZS90 particle size and potential measuring instrument. The particle size distribution diagram is shown in Figure 1, and the average particle size, PDI and Zate potential (25°C, n=10) are shown in Table 2.

Table 2 Particle size, PDI and Zate potential of blank microemulsion and paclitaxel microemulsion for injection (25°C, n=10)

composition Blank-ME PTX-ME Size(nm) 49.4±0.93 50.1±1.21 PDI 0.145±0.006 0.153±0.005 Zetapotential (mv) -0.537±0.12 -0.832±0.28

The results showed that the particle size of the two microemulsions was uniform, both about 50nm. Zate potential results showed that both microemulsions were electrically neutral.

Transmission electron microscope observation:

Take 1 mL of paclitaxel microemulsion for injection prepared in Example 1 and add it into a 10 mL volumetric flask, dilute to the mark with distilled water, and shake well. Take a drop on the copper grid, counterstain, and observe with electron microscope. The scanning image of transmission electron microscope is shown in Figure 2. The measured drug particle size is about 50nm, and its shape is spherical or quasi-spherical.

Investigation of plasma stability:

Fetal bovine serum (FBS) was diluted to a mass concentration of 10% with PBS (pH 7.4) to simulate the blood environment in vivo, and 0.5 mL of PTX-ME obtained in Example 1 and Example 2 was added to 1 mL of a mass concentration of 10% In the FBS solution, put it on a constant temperature water-bath shaker at 37°C, shake at a speed of 100rpm, and shake it for 0h, 4h, 8h, 12h, 24h, and 48h respectively. table 3.

Table 3 plasma stability investigation result (n=3)

The experimental results showed that the PTX-ME loaded with 1.6mg/g and 2.6mg/g had no significant difference in particle size between 0h and 48h, indicating that the prepared paclitaxel injection microemulsion remained stable in plasma within 48h.

Study on the storage stability of diluted PTX-ME prepared in different solvents at 4°C:

Dilute the PTX-ME obtained in Example 1 with water for injection, normal saline, and glucose for injection as solvents, place at 4°C within 0, 1, 2, 3, 4, and 5 days, observe the drug precipitation every day, and Sampling was carried out to determine the particle size, and the results are shown in Table 4.

Table 4 The particle size (nm) of diluted PTX-ME prepared in different solvents and stored at 4°C for 5 days

The experimental results showed that after the three solvents were placed for 5 days, the particle size did not change much, and there was no obvious difference. The naked eye observation was clear and transparent, and no drug precipitation was seen, indicating that the drug remained stable in the three solvents, and the drug was not precipitated.

test case 2

1. In vitro release test:

和游离紫杉醇(Free-PTX)分别配制成含紫杉醇8mg/5mL的溶液，分别取1mL滴入透析袋中(12000Da)，将透析袋密封，置于150mL含有1.0％Tween80的PBS中(pH为7.4)，置于转速为100rpm，温度为37℃的恒温振荡器中，在预定的时间点0、0.5、1、2、4、6、8、10、12，24，36，48，60，72h分别取样1mL，并补充等体积新鲜的释放介质，取样时间可根据结果进行调整，将不同时间点的样品过0.22μm的滤膜，通过HPLC检测药物浓度并计算累积释放量。紫杉醇注射用微乳(PTX-ME)、紫杉醇注射液

和游离紫杉醇(Free-PTX)的体外释放结果如图3所示。Adopt dialysis, the paclitaxel injection microemulsion (PTX-ME) that embodiment 1 makes, commercially available preparation paclitaxel injection

and free paclitaxel (Free-PTX) were prepared into a solution containing paclitaxel 8mg/5mL, respectively, 1mL was dripped into a dialysis bag (12000Da), the dialysis bag was sealed, and placed in 150mL of PBS containing 1.0% Tween80 (pH 7.4 ), placed in a constant temperature oscillator with a rotation speed of 100rpm and a temperature of 37°C, at predetermined time points 0, 0.5, 1, 2, 4, 6, 8, 10, 12, 24, 36, 48, 60, 72h Sampling 1mL respectively, and adding an equal volume of fresh release medium, the sampling time can be adjusted according to the results, the samples at different time points were passed through a 0.22μm filter membrane, the drug concentration was detected by HPLC and the cumulative release amount was calculated. Paclitaxel Microemulsion for Injection (PTX-ME), Paclitaxel Injection

and free paclitaxel (Free-PTX) in vitro release results are shown in Figure 3.

呈现相似的释放趋势，释放较为缓慢。The result shows that paclitaxel microemulsion for injection of the present invention and commercially available preparation

It showed a similar release trend, and the release was slower.

2. In vitro anti-cell proliferation experiment:

对MCF-7和MDA-MB-231细胞的细胞毒作用。将MCF-7和MDA-MB-231细胞接种于96孔细胞培养板中，细胞密度为1×10⁴个/well(180μL)，孵育24h，使细胞贴壁生长。将系列浓度的游离紫杉醇组(FreePTX)、空白微乳组、紫杉醇注射用微乳组、Taxol-Blan k组(Cremophor EL/Ethanol)、泰素组

各20μL分别加入到细胞培养孔中，使MCF-7和MDA-MB-231两种细胞中各实验组的PTX终浓度的梯度分别为：1、5、10、20、32、50μg/mL，每个浓度设置6个复孔，将两种细胞的培养板置于培养箱中孵育，孵育24h和48h两个时间点。给药结束后，弃上清，PBS洗涤2遍，每孔加入20μL的MTT溶液(5.0mg/mL)和100μL的无血清培养基，继续孵育4h。待生成紫色甲瓒结晶后，弃去上清，每孔加入200μL的DMSO。室温下水平振摇20min，使生成的结晶完全溶解。使用酶标仪于490nm波长下测定每孔吸光度值(OD值)，抗增殖活性以细胞活力来表达，计算公式如式1所示：Adopt MTT method, investigate the paclitaxel injection microemulsion of embodiment 1 and

Cytotoxicity against MCF-7 and MDA-MB-231 cells. MCF-7 and MDA-MB-231 cells were inoculated in a 96-well cell culture plate at a cell density of 1×10 ⁴ cells/well (180 μL), and incubated for 24 hours to allow the cells to grow adherently. Free paclitaxel group (FreePTX), blank microemulsion group, paclitaxel injection microemulsion group, Taxol-Blan k group (Cremophor EL/Ethanol), taxol group

Each 20 μL was added to the cell culture wells, so that the gradients of the final concentration of PTX in each experimental group in MCF-7 and MDA-MB-231 cells were: 1, 5, 10, 20, 32, 50 μg/mL, Set up 6 duplicate wells for each concentration, and place the culture plates of the two kinds of cells in an incubator for incubation at two time points of 24h and 48h. After the administration, the supernatant was discarded, washed twice with PBS, 20 μL of MTT solution (5.0 mg/mL) and 100 μL of serum-free medium were added to each well, and the incubation was continued for 4 h. After the formation of purple formazan crystals, the supernatant was discarded, and 200 μL of DMSO was added to each well. Shake horizontally at room temperature for 20 minutes to completely dissolve the generated crystals. Use a microplate reader to measure the absorbance value (OD value) of each hole at a wavelength of 490nm, and the anti-proliferation activity is expressed in terms of cell viability, and the calculation formula is as shown in formula 1:

In Formula 1, ODd is the OD value of cells incubated with preparation samples, ODb is the OD value of cell culture medium, and ODr is the OD value of cells incubated with culture medium alone.

EL/Ethanol>Blank-ME>FreePTX，制备的PTX-ME和市售的

具有相同的肿瘤抑制效果，但是空白材料(空白微乳组)相比

(Taxol-Blank组)具有更低的毒性；制剂浓度为50μg/mL孵育48h时，两种细胞存活率均小于10％。The results of the anti-cell proliferation experiment in vitro are shown in Fig. 4 . The results show that under the same concentration at the same time, the inhibition rate of each preparation on breast cancer cells is:

EL/Ethanol>Blank-ME>FreePTX, prepared PTX-ME and commercially available

Have the same tumor suppressive effect, but blank material (blank microemulsion group) compares

(Taxol-Blank group) has lower toxicity; when the concentration of the preparation is 50 μg/mL and incubated for 48 hours, the survival rate of both cells is less than 10%.

3. Tumor growth inhibition experiment:

剂量10mg/kg；(3)生理盐水(saline)。给药时间以肿瘤大小生长至约100mm³时开始计算，每三天给药，每次剂量同第一次。观察荷瘤小鼠的生活状态，于给药当天称量其体重，并用游标卡尺测量其肿瘤体积，肿瘤体积用V(mm³)表示。实验结束后，以每组荷瘤裸鼠肿瘤体积的平均值为纵坐标、测量时间为横坐标绘制肿瘤生长曲线；肿瘤生长抑制实验结果如图5所示。统计分析肿瘤体积，计算肿瘤体积抑制率。肿瘤体积抑制率的计算公式如式2所示：Take 30 BALB/c nude mice and establish the tumor-bearing nude mouse model. Seven days after inoculation with MDA-MB-231 cells, when the tumor volume grows to about 100 mm ³ , the tumor-bearing nude mice are randomly divided into 3 groups, each group 10 only. The following three groups of preparations were given by tail vein injection respectively: (1) PTX-ME dose 10mg/kg;

Dose 10mg/kg; (3) Physiological saline (saline). The administration time starts to be calculated when the tumor grows to about 100 mm ³ , and is administered every three days, and each dose is the same as the first dose. The living conditions of the tumor-bearing mice were observed, their body weight was weighed on the day of administration, and their tumor volume was measured with a vernier caliper, and the tumor volume was represented by V (mm ³ ). After the experiment, the tumor growth curve was drawn with the average tumor volume of the tumor-bearing nude mice in each group as the ordinate and the measurement time as the abscissa; the results of the tumor growth inhibition experiment are shown in FIG. 5 . The tumor volume was statistically analyzed, and the tumor volume inhibition rate was calculated. The calculation formula of tumor volume inhibition rate is shown in formula 2:

对肿瘤均有一定的抑制作用，其中PTX-ME组肿瘤抑制效果最好，PTX-ME组对肿瘤的体积抑制率为68.09％，

组对肿瘤体积抑制率为56.50％。The results showed that the administration group PTX-ME and

All have a certain inhibitory effect on tumors, among which the PTX-ME group has the best tumor inhibitory effect, and the tumor volume inhibition rate of the PTX-ME group is 68.09%.

The tumor volume inhibition rate of the group was 56.50%.

4. Tissue distribution experiment of tumor-bearing nude mice:

于给药后1h和24h处死裸鼠，分离出心、肝、脾、肺、肾、脑、肿瘤组织，样品处理后，LC-MS进样分析，荷瘤裸鼠组织分布实验结果如图6所示。After the tumor growth inhibition experiment was over, paclitaxel microemulsion and paclitaxel microemulsion and

The nude mice were killed 1h and 24h after the administration, and the heart, liver, spleen, lung, kidney, brain, and tumor tissues were separated. After the samples were processed, LC-MS was injected for analysis. The results of the tumor-bearing nude mouse tissue distribution experiment are shown in Figure 6 shown.

相比，给药1h后，在各组织和肿瘤中分布较少，给药24h后，PTX-ME在肿瘤组织浓度更高，可能是由于微乳具有靶向性。PTX-ME较

有更少的心、肝、脾、肺、肾、脑分布，更高的肿瘤分布，意味着具有更少的毒性和更好的抗肿瘤效果。The experimental results show that PTX-ME and

In contrast, after 1 hour of administration, the distribution of PTX-ME in various tissues and tumors was less, and after 24 hours of administration, the concentration of PTX-ME in tumor tissues was higher, which may be due to the targeting of microemulsions. Compared with PTX-ME

There are fewer heart, liver, spleen, lung, kidney, and brain distributions, and higher tumor distributions, which means less toxicity and better anti-tumor effects.

Although the foregoing embodiments have described the present invention in detail, it is only a part of the embodiments of the present invention, rather than all embodiments. People can also obtain other embodiments according to the embodiments of the present invention without creative work. The embodiments all belong to the protection scope of the present invention.

Claims (10)

1. A microemulsion for paclitaxel injection, characterized in that it comprises the following components in parts by weight: 4 to 8 parts of paclitaxel, 235 to 300 parts of oil phase, 210 to 275 parts of surfactant, and 70 to 90 parts of cosurfactant Parts, 1900-2500 parts of solvent for injection. 2. paclitaxel injection microemulsion according to claim 1, is characterized in that, described oily phase comprises castor oil, oleic acid, ethyl oleate, medium chain fatty acid glyceride, isopropyl myristate and vitamin E One or more of acetates. 3. paclitaxel injection microemulsion according to claim 1, is characterized in that, described tensio-active agent comprises one in Tween 80, macrogol stearate, poloxamer 188 and soybean lecithin one or more species. 4. The paclitaxel microemulsion for injection according to claim 1, wherein the co-surfactant comprises one or more of dehydrated alcohol, polyethylene glycol 400 and diethylene glycol monoethyl ether. 5. The microemulsion of paclitaxel for injection according to claim 1, wherein the solvent for injection comprises one or more of water for injection, physiological saline for injection and glucose solution for injection. 6 . The paclitaxel injection microemulsion according to any one of claims 1 to 5 , characterized in that the particle size of the paclitaxel injection microemulsion is 40 to 80 nm. 7. The microemulsion for paclitaxel injection according to any one of claims 1-5, characterized in that the pH value of the microemulsion for paclitaxel injection is 4-6. 8. The preparation method of paclitaxel microemulsion for injection according to any one of claims 1 to 7, characterized in that it comprises the following steps: The paclitaxel, some surfactants and co-surfactants are first mixed to obtain a mixed solution; The mixed solution, the oil phase, the remaining part of the surfactant and the solvent for injection are mixed for the second time, and then filtered to obtain the microemulsion of paclitaxel for injection. 9. The preparation method according to claim 8, characterized in that, the time for the first mixing is 5-15 minutes. 10. A microemulsion lyophilized powder for paclitaxel injection, characterized in that the paclitaxel injection microemulsion obtained by the microemulsion for injection of paclitaxel according to any one of claims 1 to 7 or the preparation method described in claim 8 or 9 The microemulsion is obtained by drying.

Images