CN102319214A - Lipid microsphere injection containing sodium demethyl cantharidate-phosphatide complex and preparation method thereof - Google Patents
Lipid microsphere injection containing sodium demethyl cantharidate-phosphatide complex and preparation method thereof Download PDFInfo
- Publication number
- CN102319214A CN102319214A CN201110267058A CN201110267058A CN102319214A CN 102319214 A CN102319214 A CN 102319214A CN 201110267058 A CN201110267058 A CN 201110267058A CN 201110267058 A CN201110267058 A CN 201110267058A CN 102319214 A CN102319214 A CN 102319214A
- Authority
- CN
- China
- Prior art keywords
- demethylate
- disodium cantharidinate
- injection
- phosphatide
- oil
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
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- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
Abstract
The invention discloses a lipid microsphere injection containing sodium demethyl cantharidate-phosphatide complex and preparation method thereof. The injection comprises the sodium demethyl cantharidate-phosphatide complex, fat-soluble medium, surfactant and other ingredients. In the sodium demethyl cantharidate-phosphatide complex, the molar ratio of sodium demethyl cantharidate to phosphatide is 1-1:10. The prepared sodium demethyl cantharidate-phosphatide complex improves the distribution of sodium demethyl cantharidate in oil-water 2-phase interfacial film and oil phase greatly, increases the entrapment efficiency of the medicine in preparation, realizes the interfacial film loading medicine, reduces the toxicity, and can carry out targeting drug release in vivo. The prepared lipid microsphere injection reduces the vascular stimulation of sodium demethyl cantharidate, improves the curative effect, and reduces toxic and side effect.
Description
Technical field
The present invention relates to a kind of lipide microsphere injection and preparation method thereof, be specifically related to a kind of demethylate disodium cantharidinate phosphatide composite liposome microsphere injection liquid and preparation method thereof that contains, belong to medical technical field.
Background technology
(Norcantharidin NCTD), is removed 1,2 methyl and is got by cantharidin the PTS cantharidin.The sodium salt that demethylate disodium cantharidinate makes for the norcantharidin direct hydrolysis, it only dissolves in minority organic solvent (like acetone, ethyl acetate) and hot water, and the dissolubility in cold water and oil is very low.Compare with cantharidin, the urinary system stimulation of demethylate disodium cantharidinate reduces greatly, has simultaneously than the more significant function of increasing leukocyte of cantharidin.Demethylate disodium cantharidinate is a CCSA, blocks the cycle in M, and influences its periodic duty speed.Be mainly used in premedicate or combined chemotherapies such as hepatocarcinoma, esophageal carcinoma, gastric cancer, carcinoma of gastric cardia clinically, can be used for leucocytes reduction and hepatitis B.
At present, demethylate disodium cantharidinate mainly is applied to clinical with tablet and demethylate disodium cantharidinate injection.Though demethylate disodium cantharidinate has obviously alleviated the toxicity of cantharidin, do not eliminate fully.The animal experiment study result shows that when dosage increased to a certain degree, pathological change appearred in kidney and liver organization.The strictness restriction of using dosage has also seriously hindered the performance of curative effect of medication.The part patient untoward reaction occurs during clinical use when intravenously administrable surpasses 20mg or surpasses 30mg oral every day every day, symptoms such as that oral untoward reaction mainly shows as is nauseating, vomiting, dizziness.Because injection is an alkaline solution, its pH value is about 8, and is therefore bigger to the blood vessel wall stimulation much larger than pH value of blood (7.35～7.45), is prone to cause chemical phlebitis; And because medicine itself has than strong and stimulating, also can cause vasospasm, blood reduces, and local relatively disodium cantharidinate concentration increases, and increases the weight of phlebitis, and if transfusion speed greater than velocity of blood flow, then phlebitic incidence rate obviously increases.Main adverse reaction shows as and is prone to cause inflammation of vein, and vein portion is red, swollen, hot, bitterly, and is rubescent along vein traveling strip, then red and swollenly becomes block if any oozing out, and limitation of activity is handled to take a turn for the better through hot packing and cured.Therefore clinical when being used for the transfusion of periphery superficial vein, the ratio that superficial phlebitis takes place is quite high, and the patient who has refuses with this medicine treatment, influence medicine normally use clinically.
Given this; Bring into play clinical efficacy better for making demethylate disodium cantharidinate; The research and development novel form; Advantage efficient, low toxicity with novel form is given full play to the demethylate disodium cantharidinate anti-tumor activity and is reduced toxic and side effects, and this undoubtedly will be significant to further promotion and the application of popularization cantharidin in clinical.
Summary of the invention
Technical problem to be solved by this invention is that the defective that overcomes prior art provides a kind of demethylate disodium cantharidinate phosphatide composite liposome microsphere injection liquid, and this demethylate disodium cantharidinate phosphatide composite liposome microsphere injection liquid has characteristics such as low blood vessel irritation and hypotoxicity.Further, the present invention provides this demethylate disodium cantharidinate phosphatide composite liposome microsphere injection liquid to get method for preparing.
Technical problem according to the invention is realized by following technical scheme.
A kind of demethylate disodium cantharidinate phosphatide composite liposome microsphere injection liquid, this injection contains demethylate disodium cantharidinate phosphatide complexes, fat-soluble medium, surfactant and water.
Described Injectio natarii norcantharidatis phosphatide complexes is composited in organic solvent by demethylate disodium cantharidinate and phospholipid.The mol ratio of described demethylate disodium cantharidinate and phospholipid is 1～1:10.Described phospholipid is selected from one or more in natural phospholipid or the synthetic phospholipid.Said natural phospholipid is lecithin, fabaceous lecithin, cholesterol, cholic acid class or chitosan; Said synthetic phospholipid is phosphatidyl glycerol ester or DSPC.Said demethylate disodium cantharidinate is cantharidin or the demethylate disodium cantharidinate that in the formulation preparation process, generated through reaction by cantharidin.Described organic solvent is ethyl acetate, oxolane, dichloromethane, chloroform, acetone, dimethyl sulfoxide, normal hexane, cyclohexane extraction, methanol, ethanol, isopropyl alcohol, n-butyl alcohol.
Described fat-soluble medium comprises oil phase and oil soluble emulsifying agent; Described oil phase comprises mineral oil, vegetable oil, animal oil or its mixture; Described oil soluble emulsifying agent is to be selected from natural material or the synthetic material one or more; Described oil phase is MCT Oil, soybean oil, safflower oil, Semen Maydis oil or its mixture preferably; Described natural material preferably is lecithin, fabaceous lecithin, cholesterol, oleic-acid, cholic acid class or chitosan; Described synthetic material preferably is phosphatidyl glycerol ester, DSPC.
Described surfactant is selected from phospholipid, tween, poloxamer, enuatrol, oleic acid, cholic acid, deoxycholic acid or its mixture.Described phospholipid is selected from lecithin, fabaceous lecithin or its mixture; Described tween is selected from polysorbas20, polysorbate40, polysorbate60, Tween 80, polysorbate85, or the arbitrary proportion mixture of above different size; Described poloxamer is a poloxamer 188.
Described demethylate disodium cantharidinate phosphatide composite liposome microsphere injection liquid can also contain isoosmotic adjusting agent, pH regulator agent and metal-chelator.
Said isoosmotic adjusting agent is a glycerol, sorbitol, mannitol, glucose or its mixture; Said pH regulator agent is a sodium hydroxide, hydrochloric acid or buffer salt or its mixture; Said metal-chelator is a disodium edetate, sodium calcium edetate or its mixture.
Demethylate disodium cantharidinate phosphatide composite liposome microsphere injection liquid of the present invention, preferred group becomes by weight percentage:
Injectio natarii norcantharidatis phosphatide complexes (in cantharidin) 0.01%～5%;
Fat-soluble medium 5%～30%;
Surfactant 0.5%～5%;
Isoosmotic adjusting agent 0.5%～5%;
PH regulator agent 0.01%～1%;
Metal-chelator 0%～1%;
All the other are water for injection.
More preferably:
Demethylate disodium cantharidinate complex (in cantharidin) 0.05 %～0.4%;
Fat-soluble medium 10%～20%;
Surfactant 0.5%～5%;
Isoosmotic adjusting agent 1%～3%;
PH regulator agent 0.01%～1%;
Metal-chelator 0.005%～0.03%;
All the other are water for injection.
Most preferably be:
Demethylate disodium cantharidinate complex (in cantharidin) 0. 2%;
Fat-soluble medium 10%;
Lecithin 1.2%;
Glycerol 2.5%;
Enuatrol 0.03%;
Poloxamer 188 0.2%;
Disodium edetate 0.02%;
All the other are water for injection;
Wherein said fat-soluble medium is MCT Oil (MCT), soybean oil, safflower oil, Semen Maydis oil or its mixture.
The invention provides a kind of method for preparing of demethylate disodium cantharidinate phosphatide composite liposome microsphere injection liquid, it is characterized in that comprising the steps:
(1) preparation of demethylate disodium cantharidinate phosphatide complexes: remove methyl disodium cantharidinate and phospholipid, it is added in the organic solvent, 25～70 ℃ of reflux 0.5～3 hour are to the solution clear; Take out solution and reduce pressure rotary evaporation, place vacuum drier or freezer dryer to carry out drying at last to remove organic solvent;
(2) with the demethylate disodium cantharidinate phosphatide complexes of recipe quantity, fat-soluble medium heated and stirred under 60～80 ℃ condition, all be dissolved in the fat-soluble medium, get oil phase until phosphatide complexes;
(3) surfactant and the isoosmotic adjusting agent with recipe quantity adds the extremely whole dissolvings of an amount of water for injection heated and stirred under 60～80 ℃ condition as water;
(4) utilize tissue mashing machine, slowly water is added oil phase or oil phase is added aqueous phase, with per minute 10,000～20,000 changes stirring 3～5 minutes, promptly gets slightly breast;
(5) thick breast is transferred in the high pressure dispersing emulsification machine, at 600～1200bar, homogenize is 3～10 times under 30 ℃～60 ℃ conditions;
(6) get above-mentioned Emulsion embedding in infusion bottle, fill nitrogen, 115 ℃ of sterilize down 30min or 121 ℃ of sterilization 15min.
Preferred method for preparing is:
(1) preparation of demethylate disodium cantharidinate phosphatide complexes: remove methyl disodium cantharidinate 2g, soybean phospholipid 6g adds ethyl acetate 100ml; 60 ℃ after compound 1 hour; Rotary evaporation is gone out ethyl acetate, more than 12 hours (20-30 ℃), promptly gets medicine carrying soybean phospholipid complex through vacuum drying; Airtight package is put into refrigerator cold-storage and is preserved;
(2) with the demethylate disodium cantharidinate phosphatide complexes and the soybean oil of recipe quantity: MCT=3:1, heated and stirred under 70 ℃ condition all is dissolved in the oil phase until phosphatide complexes;
(3) glycerol, enuatrol, Tween 80 and the disodium edetate with recipe quantity adds the extremely whole dissolvings of an amount of water for injection heated and stirred under 70 ℃ condition as water;
(4) oil phase is added to aqueous phase, places high-speed tissue mashing machine 1,0000～2,0000rmp stirred 3-5 minute;
(5) with 0.1 molL
-1Sodium hydroxide or hydrochloric acid solution are regulated pH value to 6.0～8.0, and the water for injection dilution is settled to recipe quantity, are transferred in the high pressure homogenizer, with 600～1000 bar pressure homogenizing 6～10 times;
(6) inflated with nitrogen, embedding.Adopt 121 ℃ of sterilizations of high-pressure rotary 15 minutes.
Methyl disodium cantharidinate phosphatide composite liposome microsphere injection liquid according to the invention is the oil-in-water submicronized emulsion that contains demethylate disodium cantharidinate that a kind of intravenous transfusion is used, and the compositions that constitutes the oil-in-water submicronized emulsion is identical with the compositions of lipide microsphere injection.
In oil phase or the biphase interfacial film of profit, distribute should be more than 80% for demethylate disodium cantharidinate in the demethylate disodium cantharidinate phosphatide composite liposome microsphere injection liquid according to the invention, and the remainder demethylate disodium cantharidinate is distributed in water and the oil phase.
Demethylate disodium cantharidinate phosphatide composite liposome microsphere injection liquid according to the invention is at first compound with demethylate disodium cantharidinate and phospholipid; To in oil, water, be written in the phospholipid layer by the medicine demethylate disodium cantharidinate of equal indissoluble; The distribution of demethylate disodium cantharidinate in biphase interfacial film of profit and oil phase improves greatly; Improved the envelop rate of medicine in preparation; Realize the interfacial film medicine carrying, improved that it is fat-soluble, improved the drug loading of medicine in preparation, reduced its toxicity and made it in vivo can targeting drug release.Then utilize phosphatide complexes to prepare Injectio natarii norcantharidatis phosphatide composite liposome microsphere injection liquid, reduced the blood vessel irritation of demethylate disodium cantharidinate, improve curative effect and reduce toxic and side effects, thereby have certain innovation property and stronger practicality.Experimentation shows: the Injectio natarii norcantharidatis lipide microsphere injection is compared with injection, and acute toxicity obviously reduces.After rat vein gives Injectio natarii norcantharidatis lipide microsphere injection and its solution-type injection; The blood medicine time graph of Injectio natarii norcantharidatis is seen Fig. 1, and the drug distribution figure of Injectio natarii norcantharidatis in each tissue of heart, liver, kidney and different time sees Fig. 2-Fig. 7.
Description of drawings
Fig. 1 NCTD blood medicine-time graph 1 Injectio natarii norcantharidatis lipide microsphere injection 2 Injectio natarii norcantharidatis solution-type injection
Fig. 2 NCTD heart Chinese medicine concentration-time graph 1 Injectio natarii norcantharidatis lipide microsphere injection 2 Injectio natarii norcantharidatis solution-type injection
Fig. 3 NCTD liver Chinese medicine concentration-time graph 1 Injectio natarii norcantharidatis lipide microsphere injection 2 Injectio natarii norcantharidatis solution-type injection
Fig. 4 NCTD kidney Chinese medicine concentration-time graph 1 Injectio natarii norcantharidatis lipide microsphere injection 2 Injectio natarii norcantharidatis solution-type injection
Respectively organize medicine scattergram 1 Injectio natarii norcantharidatis lipide microsphere injection 2 Injectio natarii norcantharidatis solution-type injection in 0.5 hour the rat body of Fig. 5 administration
Respectively organize medicine scattergram 1 Injectio natarii norcantharidatis lipide microsphere injection 2 Injectio natarii norcantharidatis solution-type injection in 6 hours the rat body of Fig. 6 administration
Respectively organize AUC value 1 Injectio natarii norcantharidatis lipide microsphere injection 2 Injectio natarii norcantharidatis solution-type injection behind Fig. 7 administration 24 h in the rat body.
The specific embodiment
Below in conjunction with the specific embodiment the present invention is described further, instance is merely provides indicative explaination, does not mean that by any way protection scope of the present invention is limited.
The preparation of embodiment 1 demethylate disodium cantharidinate raw material
Though demethylate disodium cantharidinate has injection, all be to make sodium-salt aqueous solution from the norcantharidin direct hydrolysis.Its crude drug does not have production and selling.And prepare Injectio natarii norcantharidatis from norcantharidin is a organic reaction the most basic, and therefore, our reference literature report is a raw material with the norcantharidin, through the hydrolysis salify, and dehydration, washing, drying makes Injectio natarii norcantharidatis.Reaction equation is following:
Synthetic method is following:
Sodium hydroxide 10-30g adds water 100mL, puts coldly after the dissolving, adds norcantharidin 10-30g while stirring, treat nearly all dissolvings after, a small amount of insoluble matter of filtering.Solution decompression is concentrated near doing, get a large amount of white powder solids, add ethanol 5-50mL, sucking filtration; Obtain white crystal, repeatedly wash, wash the pH value of back inspection eluate, when treating that the eluate pH value is 4-9 at every turn with ethanol; Sucking filtration, vacuum drying gets white crystalline powder.
Refining: with above-mentioned white powder 5-50g, be dissolved in the 5-50mL water, filter with the G3 sintered filter funnel, filtrating adds 100-1000mL ethanol, places sucking filtration after 2-20 hour, and vacuum drying gets the white powder solid.
The preparation of embodiment 2 phosphatide complexes
[preparation 1] demethylate disodium cantharidinate-Ovum Gallus domesticus Flavus lecithin complex
Remove methyl disodium cantharidinate (in norcantharidin) 4g, Ovum Gallus domesticus Flavus lecithin 6g adds acetone 50ml; 60 ℃ after compound 1 hour; Rotary evaporation is gone out acetone, more than 12 hours (20-30 ℃), promptly gets medicine carrying Ovum Gallus domesticus Flavus lecithin complex through vacuum drying; Airtight package is put into refrigerator cold-storage and is preserved.
[preparation 2] demethylate disodium cantharidinate-soybean lecithin complex
Remove methyl disodium cantharidinate (in norcantharidin) 2g, soybean phospholipid 6g adds ethyl acetate 100ml; 60 ℃ after compound 1 hour; Rotary evaporation is gone out ethyl acetate, more than 12 hours (20-30 ℃), promptly gets medicine carrying soybean phospholipid complex through vacuum drying; Airtight package is put into refrigerator cold-storage and is preserved.
[preparation 3] demethylate disodium cantharidinate-cholesterol complex
Remove methyl disodium cantharidinate (in norcantharidin) 1g, cholesterol 3g adds ethyl acetate 100 ml; 60 ℃ after compound 3 hours; Rotary evaporation is gone out ethyl acetate, more than 12 hours (20-30 ℃), promptly gets medicine carrying cholesterol complex through vacuum drying; Airtight package is put into refrigerator cold-storage and is preserved.
[preparation 4] demethylate disodium cantharidinate-phosphatidyl glycerol ester complexes
Remove methyl disodium cantharidinate (in norcantharidin) 0.5g, phosphatidyl glycerol ester 2g adds acetone 50 ml; 60 ℃ after compound 1 hour; Rotary evaporation is gone out acetone, more than 12 hours (20-30 ℃), promptly gets medicine carrying phosphatidyl glycerol ester complexes through vacuum drying; Airtight package is put into refrigerator cold-storage and is preserved.
[preparation 5] demethylate disodium cantharidinate-DSPC complex
Remove methyl disodium cantharidinate (in norcantharidin) 0.5g, DSPC 2g adds ethyl acetate 100 ml; 60 ℃ after compound 1 hour; Rotary evaporation goes out ethyl acetate, more than 12 hours (20-30 ℃), promptly gets medicine carrying DSPC complex through vacuum drying; Airtight package is put into refrigerator cold-storage and is preserved.
[preparation 6] demethylate disodium cantharidinate-two Semen Myristicae phosphatidyl choline complex
Remove methyl disodium cantharidinate 2g, two Semen Myristicae phosphatidyl choline 6g add oxolane 250 ml; 60 ℃ after compound 1 hour; Rotary evaporation goes out oxolane, more than 12 hours (20-30 ℃), promptly gets medicine carrying two Semen Myristicae phosphatidyl choline complex through vacuum drying; Airtight package is put into refrigerator cold-storage and is preserved.
Embodiment 3 preparation demethylate disodium cantharidinate phosphatide composite liposome microsphere injection liquids
[prescription 1] demethylate disodium cantharidinate phosphatide composite liposome microsphere injection liquid (specification: 0.5mg/ml)
| The demethylate disodium cantharidinate complex | (in cantharidin) 0.05 |
| Soybean oil | |
| 10% | |
| Lecithin | 1.2% |
| Glycerol | 2.5% |
| Enuatrol | 0.03% |
| Poloxamer 188 | 0.2% |
| Calcium disodium edetate | 0.02% |
| Water for injection | Add to 100% |
Method for preparing:
(1) glycerol for injection, poloxamer 188, enuatrol and calcium disodium edetate are scattered in an amount of water for injection, are heated to 70 ℃ to the magnetic stirring apparatus to be stirred to whole dissolvings as water;
(2) phosphatide complexes, injection lecithin in [preparation 4] are joined in the injection soybean oil, be heated to be stirred under 80 ℃ and dissolve fully as oil phase;
(3) oil phase is added to aqueous phase, places 1,8000 rmp of high-speed tissue mashing machine to stir 3-5 minute;
(4) with 0.1 molL
-1Sodium hydroxide or hydrochloric acid solution are regulated pH value to 7.5, and the water for injection dilution is settled to recipe quantity, are transferred in the high pressure homogenizer, under 40 ℃, with 800 bar pressure homogenizing 6～10 times;
(5) bottling, envelope jar nitrogen, 121 ℃ of sterilization 15 min, ice-water bath cooling rapidly promptly gets.
[prescription 2] demethylate disodium cantharidinate phosphatide composite liposome microsphere injection liquid (specification: 0.5mg/ml)
| The demethylate disodium cantharidinate complex | (in cantharidin) 0.05 % |
| Oil phase (soybean oil: MCT=3:1) | 10% |
| Glycerol | 2.5% |
| Sodium deoxycholate | 0.03% |
| Poloxamer 188 | 0.2% |
| Calcium disodium edetate | 0.02% |
| Water for injection | Add to 100% |
Method for preparing:
(1) glycerol for injection, poloxamer 188, sodium deoxycholate, calcium disodium edetate are scattered in an amount of water for injection, are heated to 70 ℃ to the magnetic stirring apparatus to be stirred to whole dissolvings as water;
(2) [preparation 2] middle phosphatide complexes is joined in the mixing oil phase of injection MCT, injection soybean oil composition, be heated to be stirred under 80 ℃ and dissolve fully as oil phase;
(3) oil phase is added to aqueous phase, places 1,8000 rmp of high-speed tissue mashing machine to stir 3-5 minute;
(4) with 0.1 molL
-1Sodium hydroxide or hydrochloric acid solution are regulated pH value to 7.5, and the water for injection dilution is settled to recipe quantity, are transferred in the high pressure homogenizer, under 40 ℃, with 800 bar pressure homogenizing 6～10 times;
(5) bottling, envelope jar nitrogen, 121 ℃ of sterilization 15 min, ice-water bath cooling rapidly promptly gets.
[prescription 3] demethylate disodium cantharidinate phosphatide composite liposome microsphere injection liquid (specification: 1mg/ml)
| The demethylate disodium cantharidinate complex | (in cantharidin) 0.1% |
| Oil phase (soybean oil: MCT=3:1) | 15% |
| Fabaceous lecithin | 2.5% |
| Glycerol | 2.5% |
| Enuatrol | 0.06% |
| Poloxamer 188 | 0.6% |
| Sodium calcium edetate | 0.02% |
| Water for injection | Add to 100% |
Method for preparing:
(1) glycerol for injection, poloxamer 188, enuatrol, sodium calcium edetate are scattered in an amount of water for injection, are heated to 80 ℃ to the magnetic stirring apparatus to be stirred to whole dissolvings as water;
(2) phosphatide complexes, injection fabaceous lecithin in [preparation 3] are joined in the mixing oil phase of injection MCT, injection soybean oil composition, be heated to be stirred under 80 ℃ and dissolve fully as oil phase;
(3) oil phase is added to aqueous phase, places 2,0000 rmp of high-speed tissue mashing machine to stir 3-5 minute;
(4) with 0.1 molL
-1Sodium hydroxide or hydrochloric acid solution are regulated pH value to 7.0, and the water for injection dilution is settled to recipe quantity, are transferred in the high pressure homogenizer, under 40 ℃, with 800 bar pressure homogenizing 6～10 times;
(5) bottling, envelope jar nitrogen, 115 ℃ of sterilization 30 min, ice-water bath cooling rapidly promptly gets.
[prescription 4] demethylate disodium cantharidinate phosphatide composite liposome microsphere injection liquid (specification: 1mg/ml)
| The demethylate disodium cantharidinate complex | (in cantharidin) 0.1% |
| Oil phase (soybean oil: MCT=1:1) | 20% |
| Lecithin | 3% |
| Mannitol | 2.5% |
| Enuatrol | 0.1% |
| Poloxamer 188 | 0.4% |
| Disodium edetate | 0.02% |
| Water for injection | Add to 100% |
Method for preparing:
(1) injection mannitol, poloxamer 188, enuatrol, disodium edetate are scattered in an amount of water for injection, are heated to 70 ℃ to the magnetic stirring apparatus to be stirred to whole dissolvings as water;
(2) phosphatide complexes, injection lecithin in [preparation 5] are joined in the mixing oil phase of injection MCT, injection soybean oil composition, be heated to be stirred under 80 ℃ and dissolve fully as oil phase;
(3) oil phase is added to aqueous phase, places 1,8000 rmp of high-speed tissue mashing machine to stir 3-5 minute;
(4) with 0.1 molL
-1Sodium hydroxide or hydrochloric acid solution are regulated pH value to 7.5, and the water for injection dilution is settled to recipe quantity, are transferred in the high pressure homogenizer, under 40 ℃, with 800 bar pressure homogenizing 6～10 times;
(5) bottling, envelope jar nitrogen, 121 ℃ of sterilization 15 min, ice-water bath cooling rapidly promptly gets.
[prescription 5] demethylate disodium cantharidinate phosphatide composite liposome microsphere injection liquid (specification: 1mg/ml)
| The demethylate disodium cantharidinate complex | (in cantharidin) 0.1% |
| Oil phase (soybean oil: MCT=1:1) | 15% |
| Lecithin | 1.8% |
| Mannitol | 2.5% |
| Enuatrol | 0.06% |
| Poloxamer 188 | 0.4% |
| Disodium edetate | 0.02% |
| Water for injection | Add to 100% |
Method for preparing:
(1) injection mannitol, poloxamer 188, enuatrol, disodium edetate are scattered in an amount of water for injection, are heated to 70 ℃ to the magnetic stirring apparatus to be stirred to whole dissolvings as water;
(2) phosphatide complexes, injection lecithin in [preparation 1] are joined in the mixing oil phase of injection MCT, injection soybean oil composition, be heated to be stirred under 80 ℃ and dissolve fully as oil phase;
(3) oil phase is added to aqueous phase, places 1,8000 rmp of high-speed tissue mashing machine to stir 3-5 minute;
(4) with 0.1 molL
-1Sodium hydroxide or hydrochloric acid solution are regulated pH value to 7.5, and the water for injection dilution is settled to recipe quantity, are transferred in the high pressure homogenizer, under 40 ℃, with 800 bar pressure homogenizing 6～10 times;
(5) bottling, envelope jar nitrogen, 121 ℃ of sterilization 15 min, ice-water bath cooling rapidly promptly gets.
[prescription 6] demethylate disodium cantharidinate phosphatide composite liposome microsphere injection liquid (specification: 2mg/ml)
| The demethylate disodium cantharidinate complex | (in cantharidin) 0.2% |
| Oil phase (soybean oil: MCT=1:3) | 20% |
| Fabaceous lecithin | 3% |
| Glycerol | 2.5% |
| Enuatrol | 0.1% |
| Poloxamer 188 | 0.8% |
| Disodium edetate | 0.05% |
| Water for injection | Add to 100% |
Method for preparing:
(1) glycerol for injection, poloxamer 188, enuatrol, disodium edetate are scattered in an amount of water for injection, are heated to 70 ℃ to the magnetic stirring apparatus to be stirred to whole dissolvings as water;
(2) phosphatide complexes, injection fabaceous lecithin in [preparation 2] are joined in the mixing oil phase of injection MCT, injection soybean oil composition, be heated to be stirred under 80 ℃ and dissolve fully as oil phase;
(3) oil phase is added to aqueous phase, places 1,8000 rmp of high-speed tissue mashing machine to stir 3-5 minute;
(4) with 0.1 molL
-1Sodium hydroxide or hydrochloric acid solution are regulated pH value to 7.5, and the water for injection dilution is settled to recipe quantity, are transferred in the high pressure homogenizer, under 40 ℃, with 800 bar pressure homogenizing 6～10 times;
(5) bottling, envelope jar nitrogen, 121 ℃ of sterilization 15 min, ice-water bath cooling rapidly promptly gets.
[prescription 7] demethylate disodium cantharidinate phosphatide composite liposome microsphere injection liquid (specification: 2mg/ml)
| The demethylate disodium cantharidinate complex | (in cantharidin) 0.2% |
| Oil phase (soybean oil: MCT=1:1) | 30% |
| Fabaceous lecithin | 4% |
| Sorbitol | 2.5% |
| Sodium deoxycholate | 0.06% |
| Poloxamer 188 | 1.0% |
| Calcium disodium edetate | 0.02% |
| Water for injection | Add to 100% |
Method for preparing:
(1) injection sorbitol, poloxamer 188, sodium deoxycholate, calcium disodium edetate are scattered in an amount of water for injection, are heated to 70 ℃ to the magnetic stirring apparatus to be stirred to whole dissolvings as water;
(2) phosphatide complexes, injection fabaceous lecithin in [preparation 6] are joined in the mixing oil phase of injection MCT, injection soybean oil composition, be heated to be stirred under 80 ℃ and dissolve fully as oil phase;
(3) oil phase is added to aqueous phase, places 2,0000 rmp of high-speed tissue mashing machine to stir 3-5 minute;
(4) with 0.1 molL
-1Sodium hydroxide or hydrochloric acid solution are regulated pH value to 7.0, and the water for injection dilution is settled to recipe quantity, are transferred in the high pressure homogenizer, under 40 ℃, with 800 bar pressure homogenizing 6～10 times;
(5) bottling, envelope jar nitrogen, 121 ℃ of sterilization 15min, ice-water bath cooling rapidly promptly gets.
[prescription 8] demethylate disodium cantharidinate phosphatide composite liposome microsphere injection liquid (specification: 4mg/ml)
| The demethylate disodium cantharidinate complex | (in cantharidin) 0.4% |
| Oil phase (soybean oil: MCT=1:1) | 30 |
| Lecithin | |
| 5% | |
| Sorbitol | 2.5% |
| Enuatrol | 0.1% |
| Poloxamer 188 | 0.4% |
| Calcium disodium edetate | 0.02% |
| Water for injection | Add to 100% |
Method for preparing:
(1) injection sorbitol, poloxamer 188, sodium deoxycholate, calcium disodium edetate are scattered in an amount of water for injection, are heated to 70 ℃ to the magnetic stirring apparatus to be stirred to whole dissolvings as water;
(2) phosphatide complexes, injection lecithin in [preparation 6] are joined in the mixing oil phase of injection MCT, injection soybean oil composition, be heated to be stirred under 80 ℃ and dissolve fully as oil phase;
(3) oil phase is added to aqueous phase, places 2,0000 rmp of high-speed tissue mashing machine to stir 3-5 minute;
(4) with 0.1 molL
-1Sodium hydroxide or hydrochloric acid solution are regulated pH value to 7.0, and the water for injection dilution is settled to recipe quantity, are transferred in the high pressure homogenizer, under 40 ℃, with 800 bar pressure homogenizing 6～10 times;
(5) bottling, envelope jar nitrogen, 121 ℃ of sterilization 15min, ice-water bath cooling rapidly promptly gets.
[prescription 9] demethylate disodium cantharidinate phosphatide composite liposome microsphere injection liquid (specification: 4mg/ml)
| The demethylate disodium cantharidinate complex | (in cantharidin) 0.4% |
| Oil phase (soybean oil: MCT=1:1) | 20% |
| Lecithin | 3% |
| Glycerol | 2.5% |
| Enuatrol | 0.06% |
| Tween 80 | 0.4% |
| Disodium edetate | 0.02% |
| Water for injection | Add to 100% |
Method for preparing:
(1) glycerol for injection, Tween 80, enuatrol, disodium edetate are scattered in an amount of water for injection, are heated to 70 ℃ to the magnetic stirring apparatus to be stirred to whole dissolvings as water;
(2) phosphatide complexes, injection lecithin in [preparation 6] are joined in the mixing oil phase of injection MCT, injection soybean oil composition, be heated to be stirred under 80 ℃ and dissolve fully as oil phase;
(3) oil phase is added to aqueous phase, places 2,0000 rmp of high-speed tissue mashing machine to stir 3-5 minute;
(4) with 0.1 molL
-1Sodium hydroxide or hydrochloric acid solution are regulated pH value to 7.5, and the water for injection dilution is settled to recipe quantity, are transferred in the high pressure homogenizer, under 40 ℃, with 800 bar pressure homogenizing 6～10 times;
(5) bottling, envelope jar nitrogen, 121 ℃ of sterilization 15min, ice-water bath cooling rapidly promptly gets.
[prescription 10] demethylate disodium cantharidinate phosphatide composite liposome microsphere injection liquid (specification: 0.5mg/ml)
| The demethylate disodium cantharidinate complex | (in cantharidin) 0.05 % |
| Oil phase (Semen Maydis oil: MCT=3:1) | 10% |
| Soybean phospholipid | 1.2% |
| Glycerol | 2.5% |
| Sodium deoxycholate | 0.03% |
| Tween 80 | 0.2% |
| Calcium disodium edetate | 0.02% |
| Water for injection | Add to 100% |
Method for preparing:
(1) glycerol for injection, Tween 80, sodium deoxycholate, calcium disodium edetate are scattered in an amount of water for injection, are heated to 70 ℃ to the magnetic stirring apparatus to be stirred to whole dissolvings as water;
(2) phosphatide complexes, injection soybean phospholipid in [preparation 1] are joined in the mixing oil phase of injection MCT, injection Semen Maydis oil composition, be heated to be stirred under 80 ℃ and dissolve fully as oil phase;
(3) oil phase is added to aqueous phase, places 1,8000 rmp of high-speed tissue mashing machine to stir 3-5 minute;
(4) with 0.1 molL
-1Sodium hydroxide or hydrochloric acid solution are regulated pH value to 7.5, and the water for injection dilution is settled to recipe quantity, are transferred in the high pressure homogenizer, under 40 ℃, with 800 bar pressure homogenizing 6～10 times;
(5) bottling, envelope jar nitrogen, 121 ℃ of sterilization 15 min, ice-water bath cooling rapidly promptly gets.
[prescription 11] demethylate disodium cantharidinate phosphatide composite liposome microsphere injection liquid (specification: 0.5mg/ml)
| The demethylate disodium cantharidinate complex | (in cantharidin) 0.05% |
| Oil phase (soybean oil: safflower oil=1:1) | 10% |
| Lecithin | 1.5% |
| Glycerol | 2.5% |
| Enuatrol | 0.03% |
| Tween 80 | 0.6% |
| Disodium edetate | 0.02% |
| Water for injection | Add to 100% |
Method for preparing:
(1) glycerol for injection, Tween 80, enuatrol, disodium edetate are scattered in an amount of water for injection, are heated to 70 ℃ to the magnetic stirring apparatus to be stirred to whole dissolvings as water;
(2) phosphatide complexes, injection lecithin in [preparation 1] are joined in the mixing oil phase of injection safflower oil, injection soybean oil composition, be heated to be stirred under 80 ℃ and dissolve fully as oil phase;
(3) oil phase is added to aqueous phase, places 2,0000 rmp of high-speed tissue mashing machine to stir 3-5 minute;
(4) with 0.1 molL
-1Sodium hydroxide or hydrochloric acid solution are regulated pH value to 7.5, and the water for injection dilution is settled to recipe quantity, are transferred in the high pressure homogenizer, under 40 ℃, with 800 bar pressure homogenizing 6～10 times;
(5) bottling, envelope jar nitrogen, 121 ℃ of sterilization 15min, ice-water bath cooling rapidly promptly gets.
[prescription 12] demethylate disodium cantharidinate phosphatide composite liposome microsphere injection liquid (specification: 0.5mg/ml)
| The demethylate disodium cantharidinate complex | (in cantharidin) 0.05% |
| Oil phase (soybean oil) | 10% |
| Lecithin | 1.2% |
| Glycerol | 2.5% |
| Enuatrol | 0.03% |
| Tween 80 | 0.2% |
| Poloxamer 188 | 0.4% |
| Disodium edetate | 0.02% |
| Water for injection | Add to 100% |
Method for preparing:
(1) glycerol for injection, Tween 80, poloxamer 188, enuatrol, disodium edetate are scattered in an amount of water for injection, are heated to 70 ℃ to the magnetic stirring apparatus to be stirred to whole dissolvings as water;
(2) phosphatide complexes, injection lecithin in [preparation 1] are joined in the injection soybean oil, be heated to be stirred under 80 ℃ and dissolve fully as oil phase;
(3) oil phase is added to aqueous phase, places 2,0000 rmp of high-speed tissue mashing machine to stir 3-5 minute;
(4) with 0.1 molL
-1Sodium hydroxide or hydrochloric acid solution are regulated pH value to 7.5, and the water for injection dilution is settled to recipe quantity, are transferred in the high pressure homogenizer, under 40 ℃, with 800 bar pressure homogenizing 6～10 times;
(5) bottling, envelope jar nitrogen, 121 ℃ of sterilization 15min, ice-water bath cooling rapidly promptly gets.
[prescription 13] demethylate disodium cantharidinate phosphatide composite liposome microsphere injection liquid (specification: 0.5mg/ml)
| The demethylate disodium cantharidinate complex | (in cantharidin) 0.05% |
| Oil phase (soybean oil) | 20% |
| Lecithin | 2.4% |
| Glycerol | 2.5% |
| Enuatrol | 0.08% |
| Tween 80 | 0.3% |
| Poloxamer 188 | 0.3% |
| Disodium edetate | 0.05% |
| Water for injection | Add to 100% |
Method for preparing:
(1) glycerol for injection, Tween 80, poloxamer 188, enuatrol, disodium edetate are scattered in an amount of water for injection, are heated to 70 ℃ to the magnetic stirring apparatus to be stirred to whole dissolvings as water;
(2) phosphatide complexes, injection lecithin in [preparation 1] are joined in the injection soybean oil, be heated to be stirred under 80 ℃ and dissolve fully as oil phase;
(3) oil phase is added to aqueous phase, places 2,0000 rmp of high-speed tissue mashing machine to stir 3-5 minute;
(4) with 0.1 molL
-1Sodium hydroxide or hydrochloric acid solution are regulated pH value to 7.5, and the water for injection dilution is settled to recipe quantity, are transferred in the high pressure homogenizer, under 40 ℃, with 800 bar pressure homogenizing 6～10 times;
(5) bottling, envelope jar nitrogen, 121 ℃ of sterilization 15min, ice-water bath cooling rapidly promptly gets.
[prescription 14] demethylate disodium cantharidinate phosphatide composite liposome microsphere injection liquid (specification: 0.5mg/ml)
| The demethylate disodium cantharidinate complex | (in cantharidin) 0.05% |
| Oil phase (Semen Maydis oil: safflower oil=1:1) | 20% |
| Soybean phospholipid | 2.4% |
| Glycerol | 2.5% |
| Sodium deoxycholate | 0.08% |
| Tween 80 | 0.3% |
| Poloxamer 188 | 0.3% |
| Disodium edetate | 0.05% |
| Water for injection | Add to 100% |
Method for preparing:
(1) glycerol for injection, Tween 80, poloxamer 188, sodium deoxycholate, disodium edetate are scattered in an amount of water for injection, are heated to 70 ℃ to the magnetic stirring apparatus to be stirred to whole dissolvings as water;
(2) phosphatide complexes, injection lecithin in [preparation 1] are joined in the mixing oil phase of injection Semen Maydis oil, injection safflower oil, be heated to be stirred under 80 ℃ and dissolve fully as oil phase;
(3) oil phase is added to aqueous phase, places 2,0000 rmp of high-speed tissue mashing machine to stir 3-5 minute;
(4) with 0.1 molL
-1Sodium hydroxide or hydrochloric acid solution are regulated pH value to 7.5, and the water for injection dilution is settled to recipe quantity, are transferred in the high pressure homogenizer, under 40 ℃, with 800 bar pressure homogenizing 6～10 times;
(5) bottling, envelope jar nitrogen, 121 ℃ of sterilization 15min, ice-water bath cooling rapidly promptly gets.
The relevant physicochemical property of the prepared demethylate disodium cantharidinate phosphatide composite liposome of prescription 1-14 microsphere injection liquid is following:
| ? | Outward appearance | Particle diameter nm | Content | Envelop rate | Related substance |
| Prescription 1 | Homogeneous is good | 178.3±45.2 | 100.3% | 85.3% | 0.27 |
| Prescription | |||||
| 2 | Homogeneous is good | 187.6±52.7 | 99.5% | 82.1% | 0.34% |
| Prescription 3 | Homogeneous is good | 173.7±43.2 | 99.1% | 83.4% | 0.25% |
| Prescription 4 | Homogeneous is good | 168.3±46.2 | 99.1% | 81.1% | 0.29 |
| Prescription | |||||
| 5 | Homogeneous is good | 162.8±43.8 | 99.2% | 86.9% | 0.37% |
| Prescription 6 | Homogeneous is good | 165.5±47.1 | 100.7% | 87.6% | 0.29% |
| Prescription 7 | Homogeneous is good | 183.1±59.8 | 98.9% | 81.9% | 0.31% |
| Prescription 8 | Homogeneous is good | 159.5±44.1 | 100.8% | 88.1% | 0.28% |
| Prescription 9 | Homogeneous is good | 168.3±35.8 | 99.4% | 83.4% | 0.29 |
| Prescription | |||||
| 10 | Homogeneous is good | 175.8±48.7 | 97.7% | 82.1% | 0.35% |
| Prescription 11 | Homogeneous is good | 157.2±57.6 | 98.6% | 86.4% | 0.33% |
| Prescription 12 | Homogeneous is good | 189.3±55.1 | 98.7% | 85.1% | 0.27% |
| Prescription 13 | Homogeneous is good | 177.5±42.8 | 100.6% | 87.0% | 0.22% |
| Prescription 14 | Homogeneous is good | 184.2±47.6 | 98.7% | 85.5% | 0.26% |
Embodiment 4 Injectio natarii norcantharidatis phosphatide composite liposome microsphere injection liquid irritation tests
(1) blood vessel irritation experiment
Select 6 of NZws for use, 3 white rabbits are in auris dextra auricular vein injection demethylate disodium cantharidinate injection Qs, and left ear is injected 5% aseptic glucose injection of same dose as contrast; 3 white rabbits are in auris dextra auricular vein injection Injectio natarii norcantharidatis phosphatide composite liposome microsphere injection liquid Qz in addition, and left ear is injected 5% aseptic glucose injection of same dose as contrast.Once a day, continuous three days, the last administration was after 24 hours, put to death white rabbit, the response situation of perusal injection site, and dissect rabbit ear blood vessel and surrounding tissue is done paraffin section, dyeing, light microscopy checking.
The result shows: two kinds of dosage form blood vessel irritation experiments of demethylate disodium cantharidinate injection perusal is the result show: the blood vessel irritation of Qz is weaker than Qs; The microscopy report shows: the Qs group has blood vessel irritation to a certain degree to NZw ear blood vessel, and the Qz group is not seen obvious irritation to NZw ear blood vessel.
(2) muscle irritation experiment
Select 4 of NZws for use, 2 of every kind of dosage forms are respectively at right lateral thigh musculus quadriceps injection Qs and Qz injection 1ml; Aseptic 5% glucose injection of left side quadriceps femoris injection equivalent is injected after 48 hours as contrast, puts to death white rabbit; Dissect and take out quadriceps femoris; Vertically cut, observe the response situation of injection site muscular tissue, confirm the order of reaction.
0 grade: no change.
1 grade: mild hyperaemia, its scope is below 0.5cm * 1.0cm.
2 grades: moderate is congested, and its scope is more than 0.5cm * 1.0cm.
3 grades: severe is congested, with myodegeneration.
4 grades: necrosis occurs, the brown degeneration is arranged.
5 grades: the popularity necrosis occurs.
Calculate 4 quadriceps femoris order of reaction summations then, test again if the difference of the peak of the quadriceps femoris order of reaction and minimum, then should be got 2 healthy rabbits in addition greater than 2.After obtaining the result, if 4 quadriceps femoris order of reaction summations, think then that the local irritation experiment of test sample is up to specification less than 10.
The result shows: the muscle irritation of Qz is weaker than Qs.
The experiment of embodiment 5 Injectio natarii norcantharidatis phosphatide composite liposome microsphere injection liquid hemolytics
Get blood 20 ml from the common carotid artery of NZw, place in the flask, stir gently, after several minutes, remove and defibrinate, take out blood, add equivalent 5% glucose injection with Glass rod, centrifugal, remove supernatant; Sedimentary erythrocyte adds 5% glucose injection again and cleans, and is centrifugal.Transparent until supernatant so repeatedly, be made into 2% suspension with 5% glucose injection by erythrocytic capacity.
Get 7 of clean tube; Numbering adds each liquid in the following table successively respectively, and the 6th pipe does not add test liquid as the blank pipe; The 7th effective distilled water replaces 5% glucose injection; Shake up, place 37 ℃ of water-baths, whether have haemolysis to take place respectively at 0.5 hour, 1 hour, 2 hours, 3 h observation.
The result shows: Qs, Qz do not see that haemolysis takes place, and the hemolytic experiment of two kinds of dosage forms is all qualified.
The experiment of embodiment 6 Injectio natarii norcantharidatis phosphatide composite liposome microsphere injection liquid anaphylaxis
Get 8 of healthy guinea pigs, each 4 of Qs, two kinds of dosage forms of Qz, every corresponding test liquid 0.5ml of guinea pig intraperitoneal injection, the next day once, totally three times.After injection first the 14th, 21 day lateral vein in the hind paw of every Cavia porcellus outside injects corresponding confession reagent liquid 1ml (dosage conversion ditto), attacks.Each intravenously administrable was observed 2 hours, as two or more person who occurs grabbing in nose, perpendicular hair, cough, the dyspnea is judged to the positive; Be judged to the positive if any one of spasm, gatism, collapse, shock, phenomena of mortality person.All are normally negative.
The result shows: Qs, two kinds of dosage forms of Qz all meet the anaphylaxis experimental standard.
Embodiment 7 Injectio natarii norcantharidatis phosphatide composite liposome microsphere injection liquid rat plasma pharmacokinetics
Dosage regimen
12 rats are divided into two groups at random, and 6 every group, one night of fasting before the experiment.The first group is a matched group, in right back vena femoralis injection demethylate disodium cantharidinate injection (2 mgmL
-1), the second group is organized for receiving examination, in right back vena femoralis injection Injectio natarii norcantharidatis phosphatide composite liposome microsphere injection liquid, and respectively at 10 min, 20 min; 30 min, 45 min, 1 h, 2 h; 4 h, 6 h, 8 h, 12 h; 24 h eye sockets are got 0.5 mL blood, put in the centrifuge tube of the sharp end of heparinization in advance, and are centrifugal, the accurate upper plasma 200 μ L that draw; Add 20 μ L waters for injection, after handling by certain method, get 20 μ L sample introductions, calculate the concentration of demethylate disodium cantharidinate in each time point sample with the standard curve on the same day.
The compartment model pharmacokinetic parameter
Average blood drug level data with 3P97 pharmacokinetics routine processes Injectio natarii norcantharidatis phosphatide composite liposome microsphere injection liquid and demethylate disodium cantharidinate injection.Judge that according to AIC and degree of fitting both models belong to, the result shows that the blood drug level data of Injectio natarii norcantharidatis phosphatide composite liposome microsphere injection liquid and demethylate disodium cantharidinate injection injection all meet two compartment models.
Non-compartment model pharmacokinetic parameter
Describe physiological disposition by blood drug level-time measured data, calculate pharmacokinetic parameter.The result shows, main pharmacokinetic parameters AUC
0-∞, AUMC
0-∞, AUC
0-t, AUMC
0-t, S
2, MRT, the VRT there was no significant difference is seen Fig. 1.
The distributed power research of embodiment 8 Injectio natarii norcantharidatis phosphatide composite liposome microsphere injection liquid rat tissues
Dosage regimen
60 rats are divided into two groups at random, 30 every group.The first group is a matched group, and in right back vena femoralis injection demethylate disodium cantharidinate injection, the second group is organized for receiving examination, in right back vena femoralis injection Injectio natarii norcantharidatis phosphatide composite liposome microsphere injection liquid, respectively at 0.5 h; 2 h, 6 h, 12 h, 24 h (every 6 rats, each 3 of first, second groups) sacrificed by decapitation animal; Take out the heart, liver, spleen, lung; Kidney, brain, clean with normal saline flushing, blot with filter paper; Take by weighing 0.5 g, add 1 mL normal saline homogenate, less than the direct homogenate of the tissue of 0.5 g, centrifugal; Get supernatant, after handling down by " tissue sample is handled and measured " item, get 20 μ L sample introductions, calculate the concentration of NCTD in each time point sample with the standard curve on the same day.
Experimental result
The meansigma methods and the time relation of each remedy,tissue's substrate concentration of each time point of laboratory animal are seen Fig. 2～7 behind laboratory animal intravenous injection Injectio natarii norcantharidatis phosphatide composite liposome microsphere injection liquid and the Injectio natarii norcantharidatis injection.
Show that by blood plasma pharmacokinetics result of the test in the rat body Injectio natarii norcantharidatis phosphatide composite liposome microsphere injection liquid does not have obvious slow releasing function in the rat body.Tissue distribution kinetics is investigated experimental result and is shown that the distribution of demethylate disodium cantharidinate in kidney, liver, heart and blood plasma is more, and in lung, spleen and brain distribution-free almost.After changing dosage form, the total distributed in liver slightly changes, do not change the distribution characteristics of other tissue in the medicine body basically.
With the demethylate disodium cantharidinate injection is reference preparation; Measure the intravital tissue distribution dynamic characteristic of rat; The result shows: after demethylate disodium cantharidinate is prepared into the administration of lipid microsphere posterior vein; Do not change medicine rat spleen, lung, kidney, brain, and blood plasma in holdup time and drug distribution amount, the distribution in hepatic tissue slightly increases.
Claims (20)
1. a demethylate disodium cantharidinate phosphatide composite liposome microsphere injection liquid is characterized in that this injection contains demethylate disodium cantharidinate phosphatide complexes, fat-soluble medium, surfactant and water.
2. demethylate disodium cantharidinate phosphatide composite liposome microsphere injection liquid according to claim 1 is characterized in that described Injectio natarii norcantharidatis phosphatide complexes is composited in organic solvent by demethylate disodium cantharidinate and phospholipid.
3. demethylate disodium cantharidinate phosphatide composite liposome microsphere injection liquid according to claim 2 is characterized in that said demethylate disodium cantharidinate is cantharidin or the demethylate disodium cantharidinate that in the formulation preparation process, generated through reaction by cantharidin; Described organic solvent is ethyl acetate, oxolane, dichloromethane, chloroform, acetone, dimethyl sulfoxide, normal hexane, cyclohexane extraction, methanol, ethanol, isopropyl alcohol, n-butyl alcohol.
4. demethylate disodium cantharidinate phosphatide composite liposome microsphere injection liquid according to claim 3, the mol ratio that it is characterized in that described demethylate disodium cantharidinate and phospholipid is 1～1:10.
5. demethylate disodium cantharidinate phosphatide composite liposome microsphere injection liquid according to claim 4 is characterized in that described phospholipid is selected from one or more in natural phospholipid or the synthetic phospholipid.
6. demethylate disodium cantharidinate phosphatide composite liposome microsphere injection liquid according to claim 5 is characterized in that said natural phospholipid is lecithin, fabaceous lecithin, cholesterol, cholic acid class or chitosan; Said synthetic phospholipid is phosphatidyl glycerol ester or DSPC.
7. demethylate disodium cantharidinate phosphatide composite liposome microsphere injection liquid according to claim 1 is characterized in that described fat-soluble medium comprises oil phase and oil soluble emulsifying agent.
8. demethylate disodium cantharidinate phosphatide composite liposome microsphere injection liquid according to claim 7 is characterized in that described oil phase comprises mineral oil, vegetable oil, animal oil or its mixture; Described oil soluble emulsifying agent is to be selected from natural material or the synthetic material one or more.
9. demethylate disodium cantharidinate phosphatide composite liposome microsphere injection liquid according to claim 8 is characterized in that described oil phase is MCT Oil, soybean oil, safflower oil, Semen Maydis oil or its mixture; Described natural material is lecithin, fabaceous lecithin, cholesterol, oleic-acid, cholic acid class or chitosan; Described synthetic material is phosphatidyl glycerol ester, DSPC.
10. demethylate disodium cantharidinate phosphatide composite liposome microsphere injection liquid according to claim 1 is characterized in that described surfactant is selected from phospholipid, tween, poloxamer, enuatrol, oleic acid, cholic acid, deoxycholic acid or its mixture.
11. demethylate disodium cantharidinate phosphatide composite liposome microsphere injection liquid according to claim 15 is characterized in that described phospholipid is selected from lecithin, fabaceous lecithin or its mixture; Described tween is selected from polysorbas20, polysorbate40, polysorbate60, Tween 80, polysorbate85, or the arbitrary proportion mixture of above different size; Described poloxamer is a poloxamer 188.
12., it is characterized in that also containing isoosmotic adjusting agent, pH regulator agent and metal-chelator according to the described demethylate disodium cantharidinate phosphatide composite liposome of arbitrary claim microsphere injection liquid among the claim 1-11.
13. demethylate disodium cantharidinate phosphatide composite liposome microsphere injection liquid according to claim 12 is characterized in that said isoosmotic adjusting agent is a glycerol, sorbitol, mannitol, glucose or its mixture; Said pH regulator agent is a sodium hydroxide, hydrochloric acid or buffer salt or its mixture; Said metal-chelator is a disodium edetate, sodium calcium edetate or its mixture.
14. demethylate disodium cantharidinate phosphatide composite liposome microsphere injection liquid according to claim 13 is characterized in that this injection contains following component by weight percentage:
Injectio natarii norcantharidatis phosphatide complexes (in cantharidin) 0.01%～5%;
Fat-soluble medium 5%～30%;
Surfactant 0.5%～5%;
Isoosmotic adjusting agent 0.5%～5%;
PH regulator agent 0.01%～1%;
Metal-chelator 0%～1%;
All the other are water for injection.
15. demethylate disodium cantharidinate phosphatide composite liposome microsphere injection liquid according to claim 14 is characterized in that this injection contains following component by weight percentage:
Demethylate disodium cantharidinate complex (in cantharidin) 0.05 %～0.4%;
Fat-soluble medium 10%～20%;
Surfactant 0.5%～5%;
Isoosmotic adjusting agent 1%～3%;
PH regulator agent 0.01%～1%;
Metal-chelator 0.005%～0.03%;
All the other are water for injection.
16. demethylate disodium cantharidinate phosphatide composite liposome microsphere injection liquid according to claim 15 is characterized in that the composition of this injection is by weight percentage:
Demethylate disodium cantharidinate complex (in cantharidin) 0. 2%;
Fat-soluble medium 10%;
Lecithin 1.2%;
Glycerol 2.5%;
Enuatrol 0.03%;
Poloxamer 188 0.2%;
Disodium edetate 0.02%;
All the other are water for injection;
Wherein said fat-soluble medium is MCT Oil (MCT), soybean oil, safflower oil, Semen Maydis oil or its mixture.
17. the lipide microsphere injection among the claim 1-16, it is characterized in that demethylate disodium cantharidinate distributes in oil phase or the biphase interfacial film of profit should be more than 80%, and the remainder demethylate disodium cantharidinate is distributed in aqueous phase.
18. the lipide microsphere injection among the claim 1-16 is characterized in that it being the oil-in-water submicronized emulsion that contains demethylate disodium cantharidinate that a kind of intravenous transfusion is used, the compositions that constitutes the oil-in-water submicronized emulsion is identical with the compositions of lipide microsphere injection.
19. the method for preparing of a demethylate disodium cantharidinate phosphatide composite liposome microsphere injection liquid is characterized in that comprising the steps:
(1) preparation of demethylate disodium cantharidinate phosphatide complexes: remove methyl disodium cantharidinate and phospholipid, it is added in the organic solvent, 25～70 ℃ of reflux 0.5～3 hour are to the solution clear; Take out solution and reduce pressure rotary evaporation, place vacuum drier or freezer dryer to carry out drying at last to remove organic solvent;
(2) with the demethylate disodium cantharidinate phosphatide complexes of recipe quantity, fat-soluble medium heated and stirred under 60～80 ℃ condition, all be dissolved in the fat-soluble medium, get oil phase until phosphatide complexes;
(3) surfactant and the isoosmotic adjusting agent with recipe quantity adds the extremely whole dissolvings of an amount of water for injection heated and stirred under 60～80 ℃ condition as water;
(4) utilize tissue mashing machine, slowly water is added oil phase or oil phase is added aqueous phase, with per minute 10,000～20,000 changes stirring 3～5 minutes, promptly gets slightly breast;
(5) thick breast is transferred in the high pressure dispersing emulsification machine, at 600～1200bar, homogenize is 3～10 times under 30 ℃～60 ℃ conditions;
(6) get above-mentioned Emulsion embedding in infusion bottle, fill nitrogen, 115 ℃ of sterilize down 30min or 121 ℃ of sterilization 15min.
20. method for preparing according to claim 19 is characterized in that comprising following steps:
(1) preparation of demethylate disodium cantharidinate phosphatide complexes: remove methyl disodium cantharidinate 2g, soybean phospholipid 6g adds ethyl acetate 100ml; 60 ℃ after compound 1 hour; Rotary evaporation is gone out ethyl acetate, more than 12 hours (20-30 ℃), promptly gets medicine carrying soybean phospholipid complex through vacuum drying; Airtight package is put into refrigerator cold-storage and is preserved;
(2) with the demethylate disodium cantharidinate phosphatide complexes and the soybean oil of recipe quantity: MCT=3:1, heated and stirred under 70 ℃ condition all is dissolved in the oil phase until phosphatide complexes;
(3) glycerol, enuatrol, Tween 80 and the disodium edetate with recipe quantity adds the extremely whole dissolvings of an amount of water for injection heated and stirred under 70 ℃ condition as water;
(4) oil phase is added to aqueous phase, places high-speed tissue mashing machine 1,0000～2,0000rmp stirred 3-5 minute;
(5) with 0.1 molL
-1Sodium hydroxide or hydrochloric acid solution are regulated pH value to 6.0～8.0, and the water for injection dilution is settled to recipe quantity, are transferred in the high pressure homogenizer, with 600～1000 bar pressure homogenizing 6～10 times;
(6) inflated with nitrogen, 121 ℃ of sterilizations of high-pressure rotary 15 minutes are adopted in embedding.
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| CN102973503A (en) * | 2012-12-25 | 2013-03-20 | 辽宁正鑫药物研究有限公司 | Norcantharidin derivative lipid microsphere injection and preparation method thereof |
| CN103655471A (en) * | 2013-12-13 | 2014-03-26 | 山东世博金都药业有限公司 | Formula of liver targeting demehylcantharidin esterified derivative submicron emulsion injection and preparation method |
| CN103720653A (en) * | 2012-10-12 | 2014-04-16 | 天津药物研究院 | Vinorelbine submicron emulsion injection and preparation method thereof |
| CN112773728A (en) * | 2020-12-31 | 2021-05-11 | 江西科技师范大学 | Compound of nano-encapsulated resveratrol, preparation method and application thereof |
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| CN115671048A (en) * | 2022-06-06 | 2023-02-03 | 沈阳信康药物研究有限公司 | Norcantharidin liposome-emulsion complex injection and preparation method thereof |
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Cited By (9)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103720653A (en) * | 2012-10-12 | 2014-04-16 | 天津药物研究院 | Vinorelbine submicron emulsion injection and preparation method thereof |
| CN103720653B (en) * | 2012-10-12 | 2016-01-20 | 天津药物研究院 | A kind of vinorelbine submicron emulsion injection and preparation method thereof |
| CN102973503A (en) * | 2012-12-25 | 2013-03-20 | 辽宁正鑫药物研究有限公司 | Norcantharidin derivative lipid microsphere injection and preparation method thereof |
| CN102973503B (en) * | 2012-12-25 | 2015-04-01 | 辽宁正鑫药物研究有限公司 | Norcantharidin derivative lipid microsphere injection and preparation method thereof |
| CN103655471A (en) * | 2013-12-13 | 2014-03-26 | 山东世博金都药业有限公司 | Formula of liver targeting demehylcantharidin esterified derivative submicron emulsion injection and preparation method |
| CN103655471B (en) * | 2013-12-13 | 2015-07-29 | 山东世博金都药业有限公司 | Liver targeting norcantharidin esterified derivatives sub-microemulsion injection and method for making |
| WO2022068585A1 (en) * | 2020-09-30 | 2022-04-07 | 北京诺康达医药科技股份有限公司 | Lipid microsphere composition and preparation method therefor |
| CN112773728A (en) * | 2020-12-31 | 2021-05-11 | 江西科技师范大学 | Compound of nano-encapsulated resveratrol, preparation method and application thereof |
| CN115671048A (en) * | 2022-06-06 | 2023-02-03 | 沈阳信康药物研究有限公司 | Norcantharidin liposome-emulsion complex injection and preparation method thereof |
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