CN1895226A - Matrine elaioplast drop ball and its preparation - Google Patents

Abstract

The invention relates to a matrine preparation and a preparation method thereof. A kind of matrine liposome drop pill is characterized in that it is mainly formed by mixing and dripping matrine liposome and auxiliary material raw materials into cooling liquid to make drop pill, and the weight of matrine liposome and auxiliary material The ratio is 1:1-10, and the auxiliary material is a water-soluble matrix; the matrine liposome is mainly prepared from matrine, soybean lecithin and cholesterol raw materials, and the mixture of matrine, soybean lecithin and cholesterol The weight ratio is 1:1-5, wherein the weight ratio of soybean lecithin and cholesterol is 1-5:1. This method solves the instability problem of matrine liposome, and adds a new oral dosage form of matrine; the matrine liposome drop pill obtained by this method has a fast disintegration speed, complete absorption, targeting, and bioavailability high degree.

Description

Matrine liposome dropping pills and preparation method thereof

technical field

The invention relates to a matrine preparation and a preparation method thereof.

Background technique

Matrine (MAT) is derived from Sophora flavescens Ait, S.alopecuroides L. and S.subprostrata Chun et T.Chen of the leguminous plant Sophora flavescens Ait. It has various pharmacological effects and clinical functions. Such as anti-tumor, anti-bacterial, anti-parasitic, anti-viral, anti-arrhythmia, swelling and diuretic, anti-allergic and alleviating leukopenia caused by cyclophosphamide. The anti-cancer mechanism of matrine shows that matrine not only has a direct killing effect on tumor cells, but also induces some tumor cells to differentiate into normal cells and promote apoptosis. In addition, while matrine is anti-tumor, it does not have a destructive effect on normal cells, and it can also increase white blood cells and improve the immune function of the body.

As a new dosage form, liposome has many unique characteristics in the application of anti-tumor therapy, such as passive targeting of tumor cells. This performance is of great significance for improving the clinical efficacy of antitumor drugs and reducing adverse reactions.

The commonly used dosage form of matrine is injection at present. Considering that the method of injection administration is complicated, it is necessary to go to the hospital for medication, which is not very convenient for patients. At the same time, the injection generally has the problem of poor stability. Preparation of matrine into liposomes can achieve targeted drug delivery. However, traditional liposomes composed of lecithin and cholesterol are thermodynamically unstable systems, and their weak stability in vivo and in vitro limits their use. greatly affect its application as a drug carrier.

Dropping pills are heated and melted by medicine and solid matrix to form a solution suspension or emulsion, and then dropped into an immiscible condensing agent. Due to the interfacial tension of the molten droplet in the condensate, it shrinks into a pill, and then condenses into a pellet. Made in solid state. Pills are a form of solid dispersion. Because it has the advantages of fast dissolution, high bioavailability, good curative effect, small side effects, good drug stability, simple preparation, and easy quality control, it has received extensive attention from the pharmaceutical industry.

Contents of the invention

The object of the present invention is to provide a kind of matrine liposome dripping pill and preparation method thereof, this method solves the instability problem of matrine liposome, has increased oral new dosage form of matrine; the matrine that this method obtains Alkaline liposome dropping pills disintegrate quickly, absorb completely, and have targeted properties.

In order to achieve the above object, the technical scheme of the present invention is: a kind of matrine liposome dripping pill, it is characterized in that it is mainly mixed and dripped into the cooling liquid by matrine liposome and auxiliary material raw material and made drop pill. Form, the weight ratio of matrine liposome and auxiliary material is 1: 1-10, and auxiliary material is water-soluble matrix; Described matrine liposome is mainly prepared from matrine, soybean lecithin and cholesterol raw material As a result, the weight ratio of matrine to soybean lecithin and cholesterol is 1:1-5, wherein the weight ratio of soybean lecithin to cholesterol is 1-5:1.

The water-soluble matrix is any one or any two or more of polyethylene glycol with a molecular weight of 4000-6000, gelatin, sodium stearate, glycerin, poloxamer with a molecular weight of 188, etc. Mixture, when any two or more are mixed, it is in any proportion.

The preparation method of above-mentioned matrine liposome dropping pill is characterized in that it comprises the steps:

1), preparation of matrine liposome: matrine liposome is mainly prepared from matrine, soybean lecithin and cholesterol raw materials, and the weight ratio of matrine, soybean lecithin and cholesterol is 1:1 -5, wherein the weight ratio of soybean lecithin and cholesterol is 1-5:1; matrine liposomes are prepared by melting method, reverse evaporation method, ether injection method or thin film ultrasonic method;

2), mixing and melting: select matrine liposomes and auxiliary materials according to the weight ratio of matrine liposomes and auxiliary materials as 1:1-10, heat to above 80°C to melt, fully mix and stir to obtain a melt , heat preservation spare;

3), dripping system: preheat the dripping pill machine, make the liquid storage room of the dropping pill machine be at a constant temperature above 80°C, put the molten material obtained above into the liquid storage room, and drop it at a dropping speed of 20-60 drops/min. Put it into the cooling liquid of the dropping pill machine to make dropping pills;

4), finished product: collect dropping pill, filter off cooling liquid, dry to obtain product.

The cooling liquid is methyl silicone oil, dimethicone oil, liquid paraffin or other vegetable oils.

The preferred 60-90°C temperature of the dropper of the dripping pill machine in the step 3); the preferred temperature of the mouth of the dripping pill machine is 70-90°C; the optimum temperature gradient of the cooling liquid is 20--10°C; The preferred range is 4-16cm.

The matrine liposome dropping pill of the invention can be used for treating diseases such as malignant tumor, arrhythmia, heart failure, viral myocarditis, coronary heart disease and the like.

The inhibitory effect of matrine liposome on mouse S ₁₈₀ , EAC, H ₂₂ tumors and the effect on immune organs were studied through pharmacological experiments. S ₁₈₀ , EAC, and H ₂₂ transplanted tumors were used to establish models, and the tumor-bearing mice in each group were dissected, and the tumor body, spleen, and thymus were taken to weigh the wet weight, calculate the tumor inhibition rate, and detect the lymphocyte transformation rate (LTT). The results showed that the dose of matrine liposome had the highest tumor inhibitory effect. Compared with the same dose of matrine, the inhibitory effects on S ₁₈₀ , EAC, and H ₂₂ were better than that of matrine, and the LTT of H ₂₂ mice was higher. in matrine. After matrine is encapsulated by liposomes, due to the targeting effect of liposomes, matrine is directionally aggregated in tumor cells and released, which has a stronger anti-tumor effect than matrine, and enhances the antitumor effect while inhibiting tumors. body immunity. Comparing the inhibitory effect of matrine liposome dropping pills and matrine liposome on mouse S ₁₈₀ , EAC, H ₂₂ tumors and the impact on immune organs by the same method, it was found that due to the matrine liposome dispersed in water In the sexual matrix, the stability of matrine liposome is increased, and its bioavailability is improved. The antitumor effect of matrine liposome drop pill is equivalent to that of matrine liposome.

The 2.2.15 cell used for in vitro culture is a good model for the screening and evaluation of anti-HBV drugs in vitro at present. Observe and compare matrine liposome dripping pills, matrine liposomes, and matrine for 2.2. 15 cells secrete HBsAg, the effect of HBeAg and the cytotoxicity of drugs, and preliminarily evaluate the anti-HBV effect of matrine liposomes. The results showed that matrine liposome dripping pills, matrine liposomes, and matrine acted on 2.2.15 cells for 11 days, and the half toxic concentrations (TC50) to cells were 8.47mg/ml and 7.29mg/ml respectively. and 1.33mg/m1, the half effective concentration (IC50) to HBsAg and HBeAg inhibition is respectively 0.065mg/m1, 0.078mg/ml, 3.35mg/ml, the therapeutic index (TI of matrine liposome to HBsAg and HBeAg) ) are 93.34, 90.46 and 2.17, respectively. It shows that matrine liposome dripping pills have a better inhibitory effect on the secretion of HBsAg and HBeAg in vitro cell culture, and can improve the anti-HBV effect of matrine.

The present invention is in the form of oral administration. Adults are orally administered three times a day, 0.1-0.2g each time, and three months is a course of treatment.

The sections of the matrine liposome dropping pills prepared by the present invention were stained with rhodan blue, and observed with an E200 Nikon ECLIPSE biological microscope to obtain that the matrine liposomes in the dropping pills had a complete form, as shown in Fig. 3 and Fig. 4 .

The matrine liposome dropping pill of the present invention not only solves the targeting property of matrine, but also solves the problem of instability of the liposome preparation because the matrine liposome is prepared as a dropping pill. At the same time, it adopts the form of liposome dropping pills, adds a new dosage form, and fills up the blank of dosage form. The preparation method of the invention is simple, and the obtained dropping pills are smooth, round and good in color, and the dosage form has fast disintegration speed, complete absorption, targeting property and high bioavailability.

Description of drawings

Fig. 1 is the picture of matrine liposome dripping pill of the present invention (batch number: 20060410)

Fig. 2 is the picture of matrine liposome dripping pill of the present invention (batch number: 20060412)

Fig. 3 is a micrograph of the cross-section of matrine liposome dropping pills of the present invention (batch number: 20060410)

Fig. 4 is a microscopic view of the section of the matrine liposome dripping pill of the present invention (batch number: 20060412)

Detailed ways

In order to better understand the present invention, the content of the present invention is further illustrated below in conjunction with the examples, but the content of the present invention is not limited to the following examples.

Example 1:

One, preparation of matrine liposomes with ether injection method

(1) prepare matrine liposome

Prepare 1% matrine solution with pH 6.8 phosphate buffer solution as solvent, take 120ml in a conical flask, dissolve 1.2g of soybean lecithin and 0.6mg of cholesterol in 60ml of ether. Slowly inject the ether solution into the matrine solution with a No. 5 needle (the needle tip is under the liquid surface) under electromagnetic stirring, stir evenly, heat to 65°C, remove the ether by volatilization, and obtain matrine liposomes.

The prepared liposome is added with a cryoprotectant and placed in a lyophilizer to freeze-dry to form a white powdery substance, which can be hydrated by adding water before use.

(2) Determination of Encapsulation Efficiency

① Separation conditions

Separation voltage: 20KV, detection wavelength: 196-400 full-wavelength scanning; measurement wavelength: 206nm Sampling method: pressure injection; injection volume: 0.5Psi×5s; buffer: 0.25M/L disodium hydrogen phosphate, diphosphate Sodium hydrogen, pH6.868; temperature 25°C.

② Preparation of reference solution

The matrine reference substance dried to constant weight was prepared into solutions of different concentrations with distilled water, and then passed through a 0.45 μm microporous membrane.

③ Preparation of the test solution

The prepared liposomes are passed through a 0.45 μm microporous membrane to be sized.

④ Preparation of Negative Control Substance Solution

Take blank liposomes and prepare according to the above method

⑤Preparation of standard curve

Take matrine reference substance 0.6mg, 1.4mg, 2mg, 4mg, 8mg, 12mg dried to constant weight, and prepare 0.3mg/ml, 0.7mg/ml, 1mg/ml, 2mg/ml, 4mg/ml with distilled water , 6mg/ml solution, through a 0.45μm microporous membrane, sample injection, high performance capillary electrophoresis (HPCE) determination. The results are as follows, see Table 1:

Table 1. Peak areas of different concentrations of matrine C (mg/ml) 0.3 0.7 1.015 2.015 4.005 6.025 A 274745 361006.5 584003.5 1058090.5 1813924 2731227.5

The concentration of matrine was used to perform linear regression on the peak area, and the regression equation and correlation coefficient were as follows: A=425787.2953C+151823.4011; r=0.99950.

The results showed that the linear relationship between concentration and peak area was good within the concentration range of 0.3-6.025mg/ml.

⑥Precision test

Take 4.005 mg/ml matrine reference substance solution and inject five times according to the above electrophoresis conditions, the peak areas are 1842036, 1815226, 1806812, 1823052, 1782494 respectively, RSD=1.21%.

(3) Stability experiment

①Heating damage:

Take matrine liposomes, heat them in a water bath at 100°C, and observe the phenomena of delamination, aggregation, and demulsification. The order of the aggregation phenomenon of liposomes prepared by various methods is ammonium sulfate gradient method, pH gradient method, Kirby method, film dispersion method, secondary emulsification method, reverse phase evaporation method, and ammonium sulfate gradient method. The liposome aggregation time of the liposome is 15min, and the time of the reverse phase evaporation method is 35min.

②Centrifugal method:

Put the liposomes in a centrifuge tube at 1000r/min to test the stratification of the liposomes. The result is that the stratification is related to the ultrasonic and stirring time involved in various methods. The longer the ultrasonic and stirring time, the better, but the ultrasonic , Stirring heat, so generally ultrasonic 15min is better, has nothing to do with the method of dispersion.

③Determination of leakage rate

Orthogonal design of four factors and three levels, after nine prescription tests, with encapsulation efficiency as an index, the best preparation conditions of ether injection method were selected. See Table 2:

Table 2. Factor level table level factor A B C D. 1 6.8 4:1 20mg 10ml 2 7.0 2:1 40mg 15ml 3 7.4 1:1 60mg 20ml

A is the pH value of PBS, B is the weight ratio of lecithin to cholesterol, C is the amount of matrine, D is the amount of water phase solvent

Table 3. Orthogonal table and experimental results serial number A B C D. Encapsulation rate (%) 1 1 1 1 1 7.66 2 1 2 2 2 35.22 3 1 3 3 3 47.37 4 2 1 2 3 11.76 5 2 2 3 1 20.95 6 2 3 1 2 11.30 7 3 1 3 2 7.75 8 3 2 1 3 50.48 9 3 3 2 1 31.61

After measuring the entrapment rate of the liposomes on the same day, put them in the refrigerator for refrigerated storage, measure the entrapment rate after 10 days, and calculate the leakage rate as follows:

Leakage rate = (encapsulation rate measured on the same day - encapsulation rate measured on a regular basis)/encapsulation rate measured on the same day × 100%. Measure the leakage rate of the liposomes prepared by the nine prescriptions of the reverse phase evaporation method to place for ten days, the results are as follows, see Table 4:

Table 4. Leakage rates of different samples sample number Encapsulation rate of the day (%) Encapsulation rate after ten days (%) Leakage rate (%) 123456789 7.6635.2247.3711.7620.9511.307.7550.4831.61 7.1233.8546.5610.9820.3311.077.1648.6230.49 7.053.891.716.632.962.047.613.683.54

It can be seen that when lecithin: cholesterol=4:1, the leakage rate of liposome is larger.

(4) Freeze-drying experiment

Liposome is not only a thermodynamically unstable system, but also a dynamically unstable system. Ordinary liposomes generally appear to deposit and darken after being placed for a month. Thus, the prepared liposomes were freeze-dried.

Freeze-drying conditions: Add trehalose to the prepared liposomes, pre-freeze until solidified, and then place them in the sample tray, first turn on the refrigerator to cool until the temperature drops to a constant temperature of -58.3°C, and then freeze at a constant temperature for 24 hours to form a white powder shape substance.

The liposomes prepared by freeze-drying were hydrated with water, and the encapsulation efficiency was measured. Its encapsulation efficiency is lower than that of the non-freeze-dried sample, which is 48.57%.

Two, prepare matrine liposome dripping pills:

Weigh 15g of polyethylene glycol (6000), put it in a beaker, heat and melt it on a water bath at 90°C, add 2g of matrine liposome, stir evenly, and keep warm for later use. Preheat the dripping pill machine to keep the temperature of the liquid storage chamber at 85±1°C, use simethicone as the coolant, pre-cool to 10°C, adjust the distance between the drip nozzle and the cooling liquid surface, pour the liquid medicine into the liquid storage chamber, Start valve, be 35 drop/min drop system with dripping speed, collect dripping pill at collecting port, remove surface coolant with gauze, vacuum-dry at normal temperature, promptly get matrine liposome dripping pill of the present invention, see Fig. 1- 2. The sections of the matrine liposome dropping pills prepared by the present invention were stained with rhodan blue, and observed with an E200 Nikon ECLIPSE biological microscope to obtain that the matrine liposomes in the dropping pills had a complete form, as shown in Fig. 3 and Fig. 4 . Here, a repeated experiment is done with a proportioning ratio, and two identical dropping pills with different production batch numbers are obtained.

Example 2:

One, reverse evaporation method prepares matrine liposome

Take 1.2g of soybean lecithin and 0.6g of cholesterol, dissolve them in 200ml of ether, and place them in a conical flask. Prepare 1% matrine solution with pH 7.4 phosphate buffer solution, under electromagnetic stirring, slowly drop 120ml of 1% matrine solution into the Erlenmeyer flask, stir to form an emulsion with ether, decompress Ether was distilled off for 1 h to obtain matrine liposomes.

Two, preparation of matrine liposome dripping pills

Weigh 10g of polyethylene glycol (6000) and 5g of polyethylene glycol (4000) into a beaker, heat and melt on a water bath at 90°C, add 3g of matrine liposome, stir evenly, and keep warm for later use. Preheat the dropping pill machine to keep the temperature of the liquid storage chamber at 90±1°C, use soybean oil as the coolant, pre-cool to 5°C, adjust the distance between the drip nozzle and the cooling liquid surface, pour the liquid medicine into the liquid storage chamber, and start the valve , be 45 drops/min drop system with dropping speed, collect dropping pill at collecting port, absorb surface coolant with gauze, vacuum-dry at normal temperature, promptly get matrine liposome dropping pill of the present invention.

Example 3:

One, film method prepares matrine liposome

Soybean lecithin 1.2g, cholesterol 0.6g, dissolve in 200ml ether, remove the organic solvent under reduced pressure with a rotary evaporator, form a uniform phospholipid film in a pear-shaped bottle, add 120ml of matrine 1% pH7.4 Phosphate buffer solution, soaking, stirring and eluting the lipid film to obtain matrine liposomes.

Two, preparation of matrine liposome dripping pills

Weigh 10 g of glycerin and 6 g of poloxamer into a beaker, heat and melt on a water bath at 88° C., add 3 g of matrine liposomes, stir evenly, and keep warm for later use. Preheat the dropping pill machine to keep the temperature of the liquid storage chamber at 88±1°C, use liquid paraffin as the coolant, pre-cool to 5°C, adjust the distance between the drip nozzle and the cooling liquid surface, pour the liquid medicine into the liquid storage chamber, and start the valve , be 60 drop/min drop system with dropping speed, collect dropping pill at collecting port, absorb surface coolant with gauze, vacuum-dry at normal temperature, promptly get matrine liposome dropping pill of the present invention.

Example 4:

One, reverse evaporation method prepares matrine liposome:

Take 0.6g of soybean lecithin and 0.6g of cholesterol, dissolve them in 200ml of ether, and place them in an Erlenmeyer flask. Prepare 1% matrine solution with pH 7.4 phosphate buffer solution, under electromagnetic stirring, slowly drop 120ml of 1% matrine solution into the Erlenmeyer flask, stir to form an emulsion with ether, decompress Ether was distilled off for 1 h to obtain matrine liposomes.

Two, prepare matrine liposome dripping pills:

Weigh 2g of gelatin, place it in a beaker, heat and melt it on a water bath at 90°C, add 2g of matrine liposome, stir evenly, and keep warm for later use. Preheat the dropping pill machine so that the temperature of the liquid storage chamber is 80±1°C, the coolant is methyl silicone oil, and it is precooled to

-10°C, adjust the distance between the drip nozzle and the cooling liquid surface, pour the liquid medicine into the liquid storage chamber, start the valve, drip at a dripping speed of 20 drops/min, collect the dripping pills at the collection port, and absorb the surface cooling with gauze agent, vacuum drying at normal temperature, to obtain matrine liposome dripping pills of the present invention.

Example 5:

One, melting method prepares matrine liposome:

Dissolve 2.5g of lecithin and 0.5g of cholesterol in 10ml of chloroform/methanol (2:1) mixed solution, evaporate the solvent in a water bath at 70°C, add 0.1g of oleic acid, polysorbate 80, sorbitan monooleate 0.5g of the mixed solution (HLB is 9.05), stirring continuously, melted; in addition, 0.6g of matrine and 0.5g of polyvinylpyrrolidone (PVP) were dissolved in an appropriate amount of newly prepared pH 7.4 phosphate buffer solution, heated to 70°C in a water bath, Then mix the two, stir at constant temperature for 1 hour, add newly prepared pH 7.4 phosphate buffer to 100ml, and sonicate for 0.5 hours. Promptly make matrine liposome.

Two, prepare matrine liposome dripping pills:

Weigh 20 g of sodium stearate, place it in a beaker, heat and melt on a water bath at 90° C., add 2 g of matrine liposome, stir evenly, and keep warm for later use. Preheat the dropping pill machine so that the liquid storage chamber has a constant temperature of 90±1°C, use dimethyl silicone oil as the coolant, pre-cool to 20°C, adjust the distance between the drip nozzle and the cooling liquid surface, pour the liquid medicine into the liquid storage chamber, Start valve, be 60 drops/min drip system with dripping speed, collect dripping pill at collecting port, suck off surface coolant with gauze, vacuum dry at normal temperature, promptly get matrine liposome dripping pill of the present invention.

Claims (5)

1. a kind of matrine liposome dripping pill is characterized in that it is mainly made drop pill and forms by matrine liposome and auxiliary material raw material mixing drop system in cooling liquid, matrine liposome and auxiliary material The weight ratio is 1:1-10, and the auxiliary material is a water-soluble matrix; the matrine liposome is mainly prepared from matrine, soybean lecithin and cholesterol raw materials, and matrine and soybean lecithin and The weight ratio of cholesterol is 1:1-5, wherein the weight ratio of soybean lecithin and cholesterol is 1-5:1. 2. a kind of matrine liposome dropping pill according to claim 1, is characterized in that: described water-soluble matrix is that molecular weight is the polyethylene glycol of 4000-6000, gelatin, sodium stearate, glycerol 1. Any one of poloxamers with a molecular weight of 188 or a mixture of any two or more, when any two or more are mixed, the ratio is arbitrary. 3. the preparation method of matrine liposome dropping pills as claimed in claim 1 is characterized in that it comprises the steps: 1), preparation of matrine liposome: matrine liposome is mainly prepared from matrine, soybean lecithin and cholesterol raw materials, and the weight ratio of matrine, soybean lecithin and cholesterol is 1:1 -5, wherein the weight ratio of soybean lecithin to cholesterol is 1-5:1; matrine liposomes are prepared by melting method, reverse evaporation method, ether injection method or thin film ultrasonic method; 2), mixing and melting: select matrine liposomes and auxiliary materials according to the weight ratio of matrine liposomes and auxiliary materials as 1:1-10, heat to above 80°C to melt, fully mix and stir to obtain a melt , heat preservation spare; 3), dripping system: preheat the dripping pill machine, make the liquid storage room of the dropping pill machine be at a constant temperature above 80°C, put the melt obtained above into the liquid storage room, and drop it at a dropping speed of 20-60 drops/min. Put it into the cooling liquid of the dropping pill machine to make dropping pills; 4), finished product: collect dropping pill, filter off cooling liquid, dry to obtain product. 4. the preparation method of a kind of matrine liposome dropping pill according to claim 3 is characterized in that: described cooling liquid is methyl silicone oil, dimethicone oil, liquid paraffin. 5. the preparation method of a kind of matrine liposome drop pill according to claim 3 is characterized in that: the temperature of the dripper of drop pill machine in described step 3) is 60-90 ℃; The temperature of the nozzle of the machine is 70-90°C; the temperature gradient of the cooling liquid is 20--10°C; the range of drop distance is 4-16cm.

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