CN100362993C - Tanshinone emulsion and its making method - Google Patents
Tanshinone emulsion and its making method Download PDFInfo
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- CN100362993C CN100362993C CNB2006100052711A CN200610005271A CN100362993C CN 100362993 C CN100362993 C CN 100362993C CN B2006100052711 A CNB2006100052711 A CN B2006100052711A CN 200610005271 A CN200610005271 A CN 200610005271A CN 100362993 C CN100362993 C CN 100362993C
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Abstract
The present invention provides tanshinone emulsion which comprises tanshinone or tanshinone IIA as active components; in addition, medicinally acceptable injection or oral-administration oil, an emulsifying agent, water for injection/water for medicine manufacture are emulsified for preparing the tanshinone emulsion. The emulsion prepared is characterized in that the distribution range of particle diameter is from 0.1 micrometre to 1 micrometre, and the average particle diameter is from 0.2 micrometre to 0.4 micrometre; the emulsion belongs to the category of sub microemulsion; and the emulsion can be taken in the modes of intravenous injection and oral administration. The emulsion can realize that the emulsion is kept stable for more than one year at room temperature, safety testing examination is qualified, and the emulsion conforms to the requirement of a clinical medicament.
Description
Technical field
The present invention relates to a kind of tanshinone emulsion and preparation method thereof, belong to drug world.
Background technology
TANSHINONES (tanshinone) is the extract of the root of salviamiltiorrhizabung (Salvia miltiorrhiza Bunge), wherein contain kind of chemical compound surplus tanshinone, Tanshinone I, cryptotanshinone, the Tanshinone II B etc. 40, because of this compounds all has o-quinone type or adjacent hydroxyl paraquinoid similar structures, majority belongs to diterpene quinone, so it is generically and collectively referred to as TANSHINONES (also claiming total tanshinone).TANSHINONES has widely effect to cardiovascular system, mainly shows as endotheliocyte protective effect, function of resisting myocardial ischemia, improves the blood metabolism and to myocardial protective effect.Its study on mechanism proves that damage has protective effect to TANSHINONES to the endotheliocyte ischemia-reperfusion, can suppress the oxidation of low density lipoprotein, LDL, suppresses the activity of lipid metabolism enzyme, improves the lipid metabolism process, for preventing and treating cardiovascular disease good action is arranged.Because TANSHINONES has cardiovascular pharmacological activity widely, therefore be applied to clinical studies and appear in the newspapers repeatly with its pharmaceutical preparation that is the main pharmacodynamics composition is developed.At present be the commercially available dosage form kind surplus in the of ten nearly of effective ingredient, relate to tablet, electuary, injection, granule, unguentum, oral liquid, injectable powder, aerosol, pill, soft capsule, drop pill, mixture etc. with the TANSHINONES.
TANSHINONES is liposoluble constituent, indissoluble in the water, and easily oxidative degradation, and instability, therefore it directly can't be prepared into can be for the injection of clinical use.Tanshinone is the main effective ingredient in the TANSHINONES, because its intestinal absorption is poor, clinical onset of action is slow, so its sulfonation is become the water solublity sodium salt by people such as money name Kun---Tanshinone I I A sodium sulfonate, successively make injection for zoopery and clinical use by Shanghai pharmacy one factory, Chinese medicine three factories etc., so far used nearly 30 years clinically, determined curative effect is sure.This shows that the active substance basis of sodium tanshinone IIA sulfate comes from tanshinone components such as Tanshinone I I A to definite therapeutical effect that cardiovascular disease had.Yet, find that through experimental study the stability of commercially available Tanshinone I I A sodium sulfonate injection is also unsatisfactory, its main effective ingredient Tanshinone I I A sodium sulfonate is also easily oxidation in aqueous solution, degraded in effect duration reaches more than 10%, is difficult to guarantee the safe and effective of preparation.Therefore, development and development of new TANSHINONES preparation have important and practical meanings.
At liposoluble constituent, Emulsion is dosage form commonly used.The dispersion of drop is very big in the Emulsion, and the performance of drug absorption and drug effect is fast, the bioavailability height.At present existing multiple medicine is prepared to Emulsion for clinical use, as emulsio olei jecoris piscis, fatsoluble vitamin Emulsion, neomycin Emulsion etc.The medicine scholar dreams of to utilize a kind of specific carrier always, and medicine directly is transported to intravital diseased region, just has medicine and leads elastic " targeted therapy ".20th century the seventies, the clinical medicine scholar of the U.S. has at first realized this imagination at laboratory, this theory of drug delivery system (DDS) has appearred in the eighties world pharmacy industry thus.Under this theory, medicine is dissolved in is scattered in water through phospholipid emulsifying in the fatty oil and makes the microsome disperse system of average particle size about 200 nanometers, be called submicron emulsion (claiming inferior nano-emulsion again).Vein is injected fast, high, the tool targeting of drug effect of back distribution with submicron emulsion.Submicron emulsion is the carrier of novel targeted medicine, alternative builds up at diseased region, and medicine is transported to the target area to greatest extent, makes medicine exceeds conventional formulation in target area concentration several times to hundreds of times, and therapeutic effect obviously improves; Medicine is few in the normal structure abundance simultaneously, and the toxic and side effects of medicine and untoward reaction meeting obviously alleviate, and reach the effect of high-efficiency low-toxicity.In addition, the submicron emulsion targeting preparation has the entrapment efficiency height, safety and good stability, characteristics such as convenient drug administration.But the physicochemical property and the clinical indication thereof of medicine depended in the use of submicron emulsion targeting preparation, is not all suitable submicron emulsion targeting preparation that is prepared into of any medicine.
Up to now, do not see the document and the patent report of any relevant TANSHINONES submicron emulsion preparation both at home and abroad as yet.Domestic only have one piece of document once the preparation and the quality evaluation of TANSHINONES microemulsion to be studied.Its mean diameter of microemulsion of (the 28th rolls up o. 11th for the preparation of TANSHINONES microemulsion and quality evaluation one Zhang Ning thereof etc., CHINA JOURNAL OF CHINESE MATERIA MEDICA, and 1081-1082 is hereinafter to be referred as documents 1) document report is 42.1nm, belongs to the nano-emulsion category.Nano-emulsion (nanoemulsion, claim microemulsion again) be that particle diameter is that the emulsion droplet of 10～100nm is dispersed in the dispersion system of colloid that forms in the another kind of liquid, it is spherical that its emulsion droplet mostly is, size is more even, transparent or semitransparent, through pressure sterilizing or centrifugally can not make it layering, belong to the thermodynamically metastable fixed system usually; And the submicron emulsion particle diameter is between 100～500nm, and outward appearance is opaque, is muddiness or emulsus, is heterogeneous disperse system, and thermodynamics and dynamics is all unstable.Therefore, it is generally acknowledged that microemulsion is stable than submicron emulsion.(Cui Fude chief editor pharmaceutics People's Health Publisher 2003 the 5th edition 356 pages)
Because ingredient complexity in the TANSHINONES under the situation that its physicochemical property still imperfectly understands, adopts the TANSHINONES microemulsion of above-mentioned documents 1 report method preparation still to have the following disadvantages:
1, the preparation of colostrum adopts 3000 watts to handle in ultrasonic 10 minutes among the microemulsion preparation technology, is difficult to realize in the big production of industry;
2, the used for intravenous injection preparation must be handled by pressure sterilizing usually, and the preparation technology of microemulsion does not carry out pressure sterilizing to the nano-emulsion after the homogenize in the documents 1, and its safety is difficult to guarantee.And for the physical and chemical stability behind its high temperature sterilize, also can't estimate, be difficult to guarantee the quality of preparation;
3, under the condition without pressure sterilizing, the stability of TANSHINONES microemulsion is also undesirable in the documents 1, place after 60 days for 25 ℃, and the visible a small amount of red granules in bottom, and envelop rate obviously descends.Above-mentioned experimental result all can not reach the preceding requirement about stability of pharmaceutical preparation listing (should keep stable more than a year at least);
4, drug loading is the important indicator in the microemulsion quality evaluation.The TANSHINONES that feeds intake altogether in every 100ml microemulsion in the prescription that documents 1 is adopted is 50mg, so its theoretic maximum drug loading is 0.05%, and the drug loading that documents 1 provides is 0.467%, exceeds nearly 10 times than its theoretical maximum drug loading.Therefore obviously, exist tangible error in data in the documents 1, for the correlational study result's of its TANSHINONES microemulsion that provides verity and accuracy, be difficult to estimate;
5, in commercially available TANSHINONES, the content of tanshinone is not from waiting (HPLC measures purity) more than 0.5%～90%, and tanshinone is the main effective ingredient in the TANSHINONES, therefore must clearly be limited the effectiveness that can guarantee preparation to the tanshinone content in the TANSHINONES raw material.Because documents 1 used content assaying method is a ultraviolet spectrophotometry, can't measure the content of single active ingredient, thereby use the tanshinone content in the TANSHINONES raw material clearly to limit to microemulsion in addition, do not carry out corresponding quality evaluation yet, can't be difficult to guarantee the effectiveness and the stability of final preparation finished product from control of quality on the raw material.
6, (it is happy etc. that the HPLC finger printing of TANSHINONES microemulsion formulation is studied a Lv Hui from another piece documents, CHINA JOURNAL OF CHINESE MATERIA MEDICA, the 29th volume o. 11th, 1047-1049, be called for short documents 2) analyze, tanshinone content is 50～62% in 10 batches of used TANSHINONES of TANSHINONES microemulsion, average out to 56.4%.Therefore, the weight percentage of tanshinone is still lower in the documents 2 employed TANSHINONES raw materials, is difficult to guarantee the effectiveness of preparation.Therefore, documents 1, the 2 described methods stability for preparing the TANSHINONES microemulsion obviously can not satisfy the clinical practice needs.Do not improve at present the relevant report of tanshinone emulsion stability yet.
Summary of the invention
Technical scheme of the present invention has provided a kind of tanshinone emulsion, and another technical scheme of the present invention has provided the preparation method of this tanshinone emulsion.
The invention provides a kind of tanshinone emulsion, it contains TANSHINONES is active component, add the emulsion liquid preparation that pharmaceutically acceptable injection or adjuvant for oral use are prepared from, wherein the particle size distribution range of interior decentralized photo is at 0.1 micron～1 micron, and mean diameter is 0.2 micron～0.4 micron.
Contained effective ingredient except that TANSHINONES, can also contain can with the effective ingredient of TANSHINONES drug combination, for example salvianolic acid B, arasaponin, ligustrazine hydrochloride, sodium ferulate etc.
Wherein, to contain 100 milligrams～1500 milligrams of TANSHINONES, injection or oil for oral use be 100g～500g, emulsifying agent 5 grams～50 grams to the every 1000ml Emulsion of tanshinone emulsion of the present invention.
Wherein, the weight percentage of tanshinone is 90-100% in the described TANSHINONES.
Wherein, described Emulsion is intravenously administrable Emulsion, Orally taken emulsion.
Oil for oral use is medicinal plant oil in the described Orally taken emulsion, as pharmaceutical grade soybean oil, Semen Maydis oil, Semen arachidis hypogaeae wet goods;
The used oil for injection of described used for intravenous injection Emulsion is generally refined vegetable oils, as soybean oil commonly used, Oleum Sesami, Oleum Camelliae etc.; Also can be long-chain, medium chain length fatty acid triglyceride and composition thereof.
Described emulsifying agent is: a kind of or its mixing in fabaceous lecithin, lecithin, Pu Luonike (F-68), Tween 80, arabic gum or the gelatin.
Further, oil for oral use is medicinal soybean oil in the described Orally taken emulsion; Its content is: containing 500 milligrams of tanshinones, soybean oil for oral use in every 1000ml Emulsion is that 250g, emulsifying agent are 15 grams, and all the other are water for pharmaceutical purposes.Wherein, described emulsifying agent is: a kind of or its mixing in fabaceous lecithin, lecithin, Pu Luonike (F-68), Tween 80, arabic gum or the gelatin.
Further, also contain glycerol for injection in described every 1000ml intravenous injection emulsion, wherein content is: containing 100 milligrams～1500 milligrams of tanshinones, oil for injection in every 1000ml Emulsion is that 100g～500g, glycerol for injection are 5 grams～50 grams, emulsifying agent 5 grams～50 grams, and all the other are water for injection.
Further, containing 500 milligrams of tanshinones, oil for injection in every 1000ml intravenously administrable Emulsion is that 200 grams, glycerol for injection are 22 grams, emulsifying agent 15 grams, and all the other are water for injection.Wherein, described emulsifying agent is: the mixture of one or more in injection fabaceous lecithin, injection lecithin, Pu Luonike (F-68) or the Tween 80.
The present invention also provides a kind of anhydrous lactitol fluid composition, it is that containing described is main component with the tanshinone emulsion, add acceptable accessories or complementary composition, the solid preparation that makes by dried newborn method preparation technologies such as freeze-drying, spray drying methods.The mean diameter of this inner decentralized photo in solid preparation water preparation back is 0.2 micron～0.4 micron.
Described adjuvant or complementary composition are: cryoprotective agent (as mannitol, glucose etc.), skeleton form agent (as lactose etc.), hydrophilic emulsifying agent and/or acetylation monoglyceride.
The present invention also provides the preparation method of tanshinone emulsion of the present invention, and wherein, the preparation method of intravenously administrable Emulsion comprises the steps:
A, take by weighing following weight proportion raw material TANSHINONES or tanshinone, adjuvant:
0.1～1.5 part of TANSHINONES or tanshinone, injection or oil for oral use are 5～50 parts of 100～500 parts, emulsifying agent;
B, the TANSHINONES of getting recipe quantity or tanshinone are dissolved in the oil phase of preheating, emulsifying agent are dissolved in the aqueous phase of preheating again;
C, above-mentioned water is changed in the tissue mashing machine, under the high-speed stirred condition, drips oil phase and go into water, be dispersed in aqueous phase to oil phase, milky or orange-yellow colostrum;
D, get above-mentioned colostrum and be transferred in the high pressure dispersing emulsification machine, homogenize to mean diameter reaches below 0.5 micron, the embedding of gained Emulsion in infusion bottle or ampoule, inflated with nitrogen, pressure sterilizing, promptly.
Particularly, the preparation method of described intravenously administrable Emulsion comprises the steps:
A, take by weighing each raw material: 100 milligrams～1500 milligrams of tanshinones, oil for injection are that 100 grams～500 grams, glycerol for injection are 5 grams～50 grams, emulsifying agent: 5 grams～50 grams, all the other are water for injection;
B, get in the oil for injection that tanshinone is dissolved in preheating, take by weighing fabaceous lecithin and glycerol again and be dissolved in an amount of water for injection that is preheated to 80 ℃;
C, above-mentioned water is changed in the tissue mashing machine, under 10000 rev/mins stirring condition, slowly drip oil phase and go into water, change stirring 10 minutes with per minute 10000, be dispersed in aqueous phase until the oil phase that is dissolved with TANSHINONES and obtain milky or orange-yellow colostrum;
D, get above-mentioned colostrum and add water for injection and make and reach full dose 1000ml, be transferred in the high pressure dispersing emulsification machine 15000Psi homogenize 3 times, the sampling and measuring particle diameter reaches below 0.5 micron to mean diameter, gets above-mentioned Emulsion embedding in infusion bottle or ampoule, inflated with nitrogen was sterilized 30 minutes down for 115 ℃.
The Emulsion of above-mentioned prepared, system mixes and the emulsifying principle according to similar, fat-soluble TANSHINONES dissolving is scattered in the oil, be prepared into Emulsion with water and emulsifying agent again, can significantly increase its dissolubility, improve its stability, can be thereby directly TANSHINONES is prepared into for oral, the intravenous injection emulsion of clinical use.Tanshinone emulsion of the present invention has solved stable problem, can at room temperature keep stable more than 1 year.
Emulsion of the present invention, particle diameter are between 0.1 micron～1 micron, and mean diameter is 200nm～400nm, belong to the submicron emulsion category, and outward appearance is opaque, is creamy white to orange-yellow.Be different from documents 1 described TANSHINONES microemulsion, must add Pu Luonike (F-68) and help emulsifying: Emulsion of the present invention only need use a small amount of phospholipid can make it keep stable as emulsifying agent, and physicochemical property is stable behind pressure sterilizing, safety testing is the result show, aseptic, the nonpyrogenic of Emulsion of the present invention's preparation, and do not have hemolytic, no anaphylaxis, nonirritant, meet the requirement of clinical application for safety and stability.
Obviously, according to foregoing of the present invention,,, can also make modification, replacement or the change of other various ways not breaking away under the above-mentioned basic fundamental thought of the present invention prerequisite according to the ordinary skill knowledge and the customary means of this area.
The specific embodiment of form is described in further detail foregoing of the present invention again by the following examples.But this should be interpreted as that the scope of the above-mentioned theme of the present invention only limits to following example.All technology that realizes based on foregoing of the present invention all belong to scope of the present invention.
Description of drawings
The particle diameter and the distribution thereof of Fig. 1 TANSHINONES vein emulsion
The Zeta potential measurement result of Fig. 2 TANSHINONES vein emulsion
The specific embodiment
The best proportion optimization test of writing out a prescription of embodiment 1 Emulsion of the present invention
Can the quality of stability of drug products be to determine it be applied to one of clinical key factor.For submicron emulsion, because it is heterogeneous disperse system, thermodynamics and dynamics is all unstable.Improve the stability of medicine carrying submicron emulsion, be a technical barrier always.All there are very big-difference in different drug loaded emulsion contained drug character and medication amount, and not having a kind of drug loaded emulsion is can known pharmaceutical emulsion prescription is conspicuous knows by inference by other.The inventor thinks, make the Emulsion for preparing stable, at first should measure physical and chemical parameters such as the dissolubility of medicine in oil, profit partition coefficient, so that determine its rational drug loading.Owing to contain kind of chemical compound surplus tanshinone, Tanshinone I, cryptotanshinone, the Tanshinone II B etc. 40 in the TANSHINONES (tanshinone), so this compounds complicated component, physicochemical property presents multiformity, is difficult to take into account various compositions and makes stable Emulsion.Therefore, should improve the purity of single active ingredient as far as possible when considering preparation TANSHINONES intravenous injection submicron emulsion, promptly the content to tanshinone clearly limits.Find that after test of many times research when the weight percentage of tanshinone reaches 90% when above in TANSHINONES, owing to the corresponding minimizing of content of other compositions, so these compositions also decrease for the influence of the stability of tanshinone emulsion.Therefore, as long as solve the influence of tanshinone, then can successfully solve the problem of tanshinone emulsion stability to emulsion stability.It is 2mg/g that the inventor has measured the dissolubility of tanshinone in the injection soybean oil first, and its partition coefficient in n-octyl alcohol/water is 2.85, and the partition coefficient in injection soybean oil/water is 2.47.This shows that tanshinone profit partition coefficient is bigger, belong to fat-soluble compound, should be prepared into stable Emulsion.(, see test example 3 about the content assaying method of tanshinone among the present invention.)
Studying the relevant physicochemical property of TANSHINONES crude drug, and then determining that we have carried out preferably the prescription of tanshinone emulsion on the basis of tanshinone content wherein (percentage by weight should greater than 90%).Adopt uniform design respectively injection oil mass, TANSHINONES (IIA) consumption, injection fabaceous lecithin amount to be investigated.Each factor level design is as follows, the results are shown in Table 1.
In the 100ml tanshinone emulsion:
Factor level
A: oil for injection (g) 5～50
B: TANSHINONES (IIA) (mg) 20～200
C: injection fabaceous lecithin (g) 0.5～5
Table 1 tanshinone emulsion prescription factor-level
| | Level | |||||||||||||
| 1 | 2 | 3 | 4 | 5 | 6 | 7 | ||||||||
| A B C | 5 20 0.5 | 10 50 1.0 | 15 80 1.5 | 20 100 2.0 | 30 120 3.0 | 40 160 4.0 | 50 200 5.0 | |||||||
Above 3 factors, 7 levels are evenly shown U by 7 levels
7(7
4) and use table to arrange test, prepare different tanshinone emulsions, with TANSHINONES accelerated test and the room temperature long-time stability that keep sample serves as to investigate index, preferred best prescription, the result shows: contain 50 milligrams of TANSHINONES and (wherein contain 47.5 milligrams of tanshinones in the every 100ml Emulsion of tanshinone emulsion of preparation, oil for injection is 20g, emulsifying agent 1.5 grams, glycerol for injection (regulating) 2.2g according to osmotic pressure, when all the other are water for injection, the gained test sample was placed 6 months for 4 ℃ at low temperature, high temperature was placed 6 months for 45 ℃, room temperature is placed after 12 months for 25 ℃, the physical appearance of sample, pH value and envelop rate have no significant change, and have good stability.Experimental result in view of the above obtains the best prescription of tanshinone emulsion of the present invention.
The preparation of embodiment 2 Emulsions of the present invention
Soybean lecithin for injection 12g and glycerol for injection 22g are inserted high-speed tissue mashing machine, add and be preheated to 80 ℃ water for injection in right amount, stir and make phospholipid be dispersed in aqueous phase; Other gets 500 milligrams of TANSHINONES (wherein containing 450 milligrams of tanshinones) and places the oil for injection of 200g preheating, after the dissolving fully oil phase, be cooled to 80 ℃ standby.Oil phase is slowly added aqueous phase, high-speed stirred 10 minutes (10000 rev/mins), make oil phase be dispersed in aqueous phase, be prepared into orange-yellow colostrum, colostrum is added injection water to 1000 milliliter, place in the high pressure dispersing emulsification machine emulsifying repeatedly 3 times, reach below 0.5 micron to mean diameter, fill the nitrogen fill again, pressure sterilizing promptly gets the TANSHINONES vein emulsion.
The preparation of embodiment 3 Emulsions of the present invention
Soybean lecithin for injection 12g, Pu Luonike (F-68) 3g and glycerol for injection 22g dissolving are scattered in the water for injection that is preheated to 80 ℃ in right amount, are prepared into homodisperse water; Other gets among the oil for injection 100g that 100 milligrams of TANSHINONES (wherein containing 100 milligrams of tanshinones) add preheating, after the dissolving fully oil phase, be cooled to 80 ℃ standby.Oil phase is slowly added aqueous phase, and high-speed stirred 10 minutes (10000 rev/mins) is prepared into faint yellow colostrum.Colostrum is added injection water to 1000 milliliter, put in the high pressure dispersing emulsification machine homogenize 3 times, reach below 0.5 micron to mean diameter, fill the nitrogen fill again, sterilization promptly makes vein emulsion.
The preparation of embodiment 4 Emulsions of the present invention
Injection soybean phospholipid 15g and glycerol for injection 22g dissolving are scattered in the water for injection that is preheated to 80 ℃ in right amount, prepare homodisperse water; Other gets among the oil for injection 200g that 500 milligrams of TANSHINONES (wherein containing 500 milligrams of tanshinones) add preheating, after the dissolving fully oil phase, be cooled to 80 ℃ standby.Oil phase is slowly added aqueous phase, and high-speed stirred 10 minutes (10000 rev/mins) is prepared into orange-yellow colostrum.Colostrum is added injection water to 1000 milliliter, put in the high pressure dispersing emulsification machine homogenize 3 times, reach below 0.5 micron to mean diameter, fill the nitrogen fill again, sterilization promptly makes vein emulsion.
The preparation of embodiment 5 Emulsions of the present invention
Injection soybean phospholipid 20g and glycerol for injection 22g dissolving are scattered in the water for injection that is preheated to 80 ℃ in right amount, prepare homodisperse water; Other gets TANSHINONES 1 gram (wherein containing tanshinone 0.95 gram) and adds among the oil for injection 300g of preheating, after the dissolving fully oil phase, be cooled to 80 ℃ standby.Oil phase is slowly added aqueous phase, and high-speed stirred 10 minutes (10000 rev/mins) is prepared into orange-yellow colostrum.Colostrum is added injection water to 1000 milliliter, put in the high pressure dispersing emulsification machine homogenize 3 times, reach below 0.5 micron to mean diameter, fill the nitrogen fill again, sterilization promptly makes vein emulsion.
The preparation of embodiment 6 Emulsions of the present invention
Injection soybean phospholipid 25g and glycerol for injection 25g dissolving are scattered in the water for injection that is preheated to 80 ℃ in right amount, prepare homodisperse water; Other gets tanshinone 1.5 gram and adds among the oil for injection 400g of preheatings, after the dissolving fully oil phase, be cooled to 80 ℃ standby.Oil phase is slowly added aqueous phase, and high-speed stirred 10 minutes (10000 rev/mins) is prepared into orange-yellow colostrum.Colostrum is added injection water to 1000 milliliter, put in the high pressure dispersing emulsification machine homogenize 3 times, reach below 0.5 micron to mean diameter, fill the nitrogen fill again, sterilization promptly makes vein emulsion.
The preparation of embodiment 7 Emulsions of the present invention
Injection soybean phospholipid 150g, glycerol for injection 220g and salvianolic acid B 15g dissolving are scattered in the water for injection that is preheated to 80 ℃ in right amount, prepare homodisperse water; Other gets TANSHINONES 5 gram (wherein containing tanshinone 4.8 grams) and adds among the oil for injection 3000g of preheating, after the dissolving fully oil phase, be cooled to 80 ℃ standby.Oil phase is slowly added aqueous phase, and high-speed stirred 10 minutes (10000 rev/mins) is prepared into orange-yellow colostrum.Colostrum is added injection water to 10 liter, put in the high pressure dispersing emulsification machine homogenize 3 times, fill the nitrogen fill again, sterilization promptly makes vein emulsion.
The preparation of embodiment 8 Emulsions of the present invention
Soybean lecithin 200g, Pu Luonike (F-68) 30g and ethyl hydroxybenzoate 10g and water for pharmaceutical purposes are made decentralized photo be preheated to 80 ℃, mix with the medicinal soybean oil of the 3000 gram preheatings that are dissolved with TANSHINONES 15 grams (wherein containing tanshinone 13.5 grams), high-speed stirred 10 minutes (10000 rev/mins), be prepared into 10 liters even Emulsion, fill the nitrogen fill again, gland promptly gets oral breast.
The preparation of embodiment 9 Emulsions of the present invention
Soybean lecithin for injection 15g and glycerol for injection 22g are inserted high-speed tissue mashing machine, add and be preheated to 80 ℃ water for injection in right amount, stir and make phospholipid be dispersed in aqueous phase; Other gets 500 milligrams of TANSHINONES (wherein containing 500 milligrams of tanshinones) and places the oil for injection of 200g preheating, after the dissolving fully oil phase, be cooled to 80 ℃ standby.Oil phase is slowly added aqueous phase, high-speed stirred 10 minutes (10000 rev/mins), make oil phase be dispersed in aqueous phase, be prepared into orange-yellow colostrum, colostrum is added injection water to 1000 milliliter, place in the high pressure dispersing emulsification machine emulsifying repeatedly 3 times, reach below 0.5 micron to mean diameter, add 80g glucose or mannitol and mix with Emulsion, extremely-45 ℃ of pre-freezes as the lyophilizing caffolding agent, put lyophilization in the freezer dryer, be prepared into the solid freeze-dried emulsion.
By the foregoing description explanation, in the range of choice of this described raw material, adjuvant, but equal preparation cost invention Emulsion.
Below prove beneficial effect of the present invention by medicine physics and chemistry of the present invention test, stability test, toxicological test.
The mensuration of 1 diameter of aspirin particle distribution of the present invention of test example and Zeta potential
Get the prepared Emulsion of the embodiment of the invention 2, after placing 1 year under room temperature (lucifuge) condition, its particle diameter and distribution thereof are measured, concrete grammar is as follows: the vein emulsion 1ml that gets the present invention's preparation, add an amount of distilled water dilution, with Malvern-3000 type particle diameter and Zeta potential analyzer (Britain), measure the particle size distribution and the Zeta potential of Emulsion, the results are shown in Figure 1～Fig. 2.
As seen from Figure 1, the particle size distribution of gained tanshinone emulsion is even, PDI=0.013, and be normal distribution.The mean diameter of vein emulsion is the 287nm (see figure 1), and Zeta potential is-the 25.8mV (see figure 2) measurement result: the particle size distribution range of interior decentralized photo is at 0.1 micron～1 micron, and mean diameter is 0.2 micron～0.4 micron.
Test example 2 medicine stabilities of the present invention detect
Reported the method and the result that the TANSHINONES microemulsion are carried out study on the stability in the documents 1, we have also adopted same procedure to investigate the stability that the present invention prepares Emulsion, and the microemulsion stability of reporting with this and documents 1 compares research.
The microemulsion of documents 1 report was placed 3 months down at 4 ℃ and 25 ℃, by the investigation of outward appearance, pH, envelop rate being checked the stability of this microemulsion, the results are shown in Table 2.
The study on the stability result of TANSHINONES microemulsion in table 2 documents
| Time (my god) | Outward appearance | pH | Envelop rate (%) | |||
| 4℃ | 25℃ | 4℃ | 25℃ | 4℃ | 25 |
|
| 0 | Homogeneous | Homogeneous | 4.58 | 4.58 | 96.59 | 96.59 |
| 10 | Homogeneous | Homogeneous | 4.56 | 4.51 | 96.47 | 96.03 |
| 20 | Homogeneous | Homogeneous | 4.38 | 4.45 | 96.41 | 95.89 |
| 30 | Homogeneous | Homogeneous | 4.61 | 4.43 | 96.35 | 94.12 |
| 60 | Homogeneous | The visible a small amount of red granules in bottom | 4.36 | 4.31 | 96.26 | 91.91 |
| 90 | Homogeneous | Bottom red color visible granule | 4.57 | 4.35 | 96.19 | 90.40 |
By table 2 as seen, the TANSHINONES microemulsion is stable not satisfactory in the documents 1, and room temperature is placed after 60 days for 25 ℃, the visible a small amount of red granules in bottom, and envelop rate obviously descends after 90 days.Above-mentioned experimental result can not reach in the new drug research technological guidance principle requirement about pharmaceutical preparation stability.
The inventor places 45 ℃ of 6 months, high temperature to the tanshinone emulsion (content of tanshinone is 90%w/w in the TANSHINONES raw material) of the embodiment of the invention 2 preparation in 4 ℃ of low temperature and places after 25 ℃ of 6 months, room temperature place 12 months, the outward appearance of test sample, pH and envelop rate, investigate the stability of this product, the results are shown in Table 3.
The study on the stability result of the tanshinone emulsion of table 3 the present invention preparation
| Time | Outward appearance | pH | TANSHINONES envelop rate (%) | Tanshinone I I A envelop rate (%) | ||||||||
| 4℃ | 25 ℃ | 45 ℃ | 4℃ | 25℃ | 45℃ | 4℃ | 25℃ | 45℃ | 4℃ | 25℃ | 45 |
|
| 0 month | Homogeneous | Homogeneous | Homogeneous | 5.22 | 5.22 | 5.22 | 97.71 | 97.71 | 97.71 | 97.54 | 97.54 | 97.54 |
| January | Homogeneous | Homogeneous | Homogeneous | 5.19 | 5.25 | 5.18 | 97.21 | 97.61 | 97.81 | 97.34 | 97.76 | 97.62 |
| February | Homogeneous | Homogeneous | Homogeneous | 5.20 | 5.22 | 5.26 | 97.64 | 97.75 | 97.69 | 97.57 | 97.65 | 97.35 |
| March | Homogeneous | Homogeneous | Homogeneous | 5.25 | 5.23 | 5.30 | 97.83 | 97.80 | 97.58 | 97.44 | 97.33 | 97.55 |
| June | Homogeneous | Homogeneous | Homogeneous | 5.24 | 5.29 | 5.23 | 97.88 | 97.81 | 97.63 | 97.71 | 97.23 | 97.31 |
| JIUYUE | — | Homogeneous | — | — | 5.17 | - | - | 97.74 | — | — | 97.64 | — |
| December | — | Homogeneous | — | — | 5.21 | - | - | 97.70 | — | — | 97.55 | — |
By table 3 as seen, adopt the emulsion stability of the present invention's preparation good.With wherein the TANSHINONES and the content of tanshinone is evaluation index, and 4 ℃ of low temperature are placed 45 ℃ of 6 months, high temperature and placed 6 months, room temperature and place after 12 months for 25 ℃ and check that envelop rate has no significant change.To sum up the result adopts the emulsion stability of the present invention's preparation obviously to be better than the microemulsion that documents 1 prepares as can be known.This Emulsion can at room temperature keep stable more than 1 year, meets in the national new drug research technological guidance principle requirement about preparation stability fully.
Test example 3 drug quality control methods of the present invention
The content assaying method of TANSHINONES among the present invention: list of references (Zhou Yaqiu etc. the assay of total tanshinone and Borneolum Syntheticum research in the compound Salviae Miltiorrhizae oral liquid. Chinese herbal medicine .1992.23 (6): 292-293) described method, adopt ultraviolet spectrophotometry, the detection wavelength is 270nm, (available from middle inspection institute) is reference substance preparation standard curve with tanshinone, measures and also calculates TANSHINONES content (TANSHINONES content is in tanshinone).
The content assaying method of tanshinone among the present invention: the employing high performance liquid chromatography (Wang Xiao waits quietly. and the content of tanshinone is relatively in the red rooted salvia of the different places of production. West China pharmaceutical journal .2004.19 (2): 142-144), with the octadecyl silane is immobile phase, methanol: water (volume ratio 80: 20) mixed solution is a mobile phase, the detection wavelength is 270nm, with tanshinone reference substance (available from middle inspection institute) preparation reference substance solution, external standard method is calculated tanshinone content.
The mensuration of test example 4 pharmaceutical emulsion envelop rates of the present invention
With the TANSHINONES is index: with reference to documents (preparation of TANSHINONES microemulsion and quality evaluation-Zhang Ning thereof etc., CHINA JOURNAL OF CHINESE MATERIA MEDICA, the 28th volume o. 11th, 1081-1082) described method, adopt the centrifugal Emulsion of supercentrifugation, 40000r/min, centrifugal 30min, get supernatant 0.5ml, dilute with dehydrated alcohol with dichloromethane dissolving back, the 270nm place surveys absorbance, according to tanshinone standard curve equation: A=0.0063C+0.0017 (r=0.9998), can determine the content M of entrapped TANSHINONES
1(TANSHINONES content is in tanshinone).The TANSHINONES total amount is M in the Emulsion
0, be calculated as follows in the tanshinone emulsion weight envelop rate Q to TANSHINONES.
Q＝M
1/M
0×100％
The result: with the TANSHINONES is index, and the prepared vein emulsion envelop rate of the present invention is all greater than 97%.
With the tanshinone is index: adopt the centrifugal Emulsion of supercentrifugation, 40000r/min, centrifugal 30min, get supernatant 0.5ml, with the dehydrated alcohol dilution,, can determine the content M of entrapped tanshinone with dichloromethane dissolving back with high effective liquid chromatography for measuring tanshinone content
1The tanshinone total amount is M in the Emulsion
0, be calculated as follows the weight envelop rate Q of tanshinone emulsion to tanshinone.
Q＝M
1/M
0×100％
The result: with the tanshinone is index, and the prepared vein emulsion envelop rate of the present invention is all greater than 97%.
Test example 5 asepsis test in medication of the present invention:
Get the microemulsion that the inventive method prepares Emulsion and prepares by documents 1 method, check according to sterility test method (2000 editions one appendix XI H of Chinese Pharmacopoeia), Emulsion sterility test with the inventive method preparation is qualified, and is defective by the microemulsion sterility test of documents 1 method preparation.
Test example 6 medicine pyrogen tests of the present invention:
Get the microemulsion that the inventive method prepares Emulsion and prepares by documents 1 method, check according to pyrogen test (2000 editions one appendix XI D of Chinese Pharmacopoeia), with the Emulsion nonpyrogenic of the inventive method preparation, defective by the microemulsion pyrogen test of documents 1 method preparation.
The 7 drug sensitivity tests of the present invention of test example:
Test method: Cavia porcellus is divided into 3 groups at random by body weight, every group 6,1,2 liang of group Cavia porcellus every other day once, 0.5ml/ sensitization of continuous three lumbar injection TANSHINONES used for intravenous injection Emulsion, 1,2 liang of group only excites respectively at 14 days behind the first time lumbar injection and 21 days digital veins of the foot injection TANSHINONES used for intravenous injection Emulsion 1.0ml/; 3 groups of Cavia porcellus every other day once, 0.5ml/ sensitization of continuous three lumbar injections, 20% egg clear liquid, 14 days digital veins of the foot after injection are injected 20% egg clear liquid 1.0ml/ and are only excited, and all observe symptoms of allergic in 15 minutes after booster injection for three groups.
Result of the test: two groups of Cavia porcelluss of TANSHINONES used for intravenous injection Emulsion are attacked with original liquid respectively at 14 days and the 21st day behind the 1st lumbar injection anaphylactic reaction are not all taken place; And positive controls (3 groups) Cavia porcellus occurs dyspnea in injecting in back 2 minutes, and tic is fallen down, and is then dead, and Cavia porcellus death all occurs in injects in back 1～3 minute.
Conclusion (of pressure testing): under this experimental condition, TANSHINONES used for intravenous injection Emulsion is not had sensitization to trying Cavia porcellus.
The 8 medicine hemolytic tests of the present invention of test example:
Test method: get 7 in test tube, 1～5 pipe adds the TANSHINONES used for intravenous injection Emulsion of 0.1ml, 0.2ml, 0.3ml, 0.4ml, 0.5ml respectively, and be diluted to 2.5ml with 10% glucose injection, add in No. 6 test tubes in 10% glucose injection 2.5ml, No. 7 test tubes and add distilled water 2.5ml (complete hemolysis contrast).Last every pipe all adds 2% rabbit erythrocyte suspension 2.5ml, shakes up gently, puts in 37 ℃ of water-baths, writes down the haemolysis and the coagulation situation of 15min, 30min, 45min, 1h, 2h, 3h, each pipe of 4h respectively.
Result of the test: TANSHINONES used for intravenous injection Emulsion 1～5 pipe did not all cause haemolysis and agglutination in 4 hours.
Conclusion (of pressure testing): under this experiment condition, TANSHINONES used for intravenous injection Emulsion does not have haemolysis and agglutination to rabbit erythrocyte.
The 9 medicine irritation tests of the present invention of test example:
Test method: get 2 of new zealand rabbits, all slowly inject TANSHINONES used for intravenous injection Emulsion 4ml/kg in left auricular vein, auris dextra edge vein is injection 10% glucose injection 4ml/kg slowly, once a day, injected continuously three days, begin for the first time in injection, observe the injection site every day and have or not edema, erythema, last was injected back 24 hours, cut ear from the basal part of the ear and put 10% formalin fixingly, did the pathology cut sections for microscopic examination then.
Result of the test: TANSHINONES used for intravenous injection Emulsion is given injection every day of rabbit ear edge vein once, and continuous three days, the generation that edema and erythema are arranged was not seen in 2 rabbit ear injection sites.Histopathology is observed, and rabbit ear epidermal structure is normal.Under the epidermis papillary layer and reticular layer do not have inflammatory cell ooze out, no hemorrhage, no thrombosis formation in the blood vessel, accessory structure is normal.
Conclusion (of pressure testing): TANSHINONES used for intravenous injection Emulsion is to rabbit ear edge vein blood vessel nonirritant.
As can be seen from the above results, adopt the Emulsion of the present invention's preparation safe and reliable, no anaphylaxis, hemolytic and zest meet the related request of clinical application.And the microemulsion that adopts documents 1 reported method to prepare, owing to there are not pressure sterilizing, safety not to meet the requirement of intravenous injection medication.
The Emulsion that adopts the inventive method preparation has been carried out anaphylaxis, hemolytic and irritation test, and result of the test shows that the emulsion stability of the present invention's preparation is good, and no anaphylaxis, hemolytic and zest meet the related request of clinical application.
Claims (7)
1. tanshinone emulsion, it is characterized in that: it is to be active component with the TANSHINONES, add the emulsion liquid preparation that pharmaceutically acceptable injection additives are prepared from, wherein the particle size distribution range of interior decentralized photo is at 0.1 micron～1 micron, mean diameter is 0.2 micron～0.4 micron, every 1000ml Emulsion contains 100 milligrams～1500 milligrams of TANSHINONES, oil for injection 100 grams～500 grams, emulsifying agent 5 grams～50 grams, described oil for injection is soybean oil or medium chain length fatty acid triglyceride, described emulsifying agent is a fabaceous lecithin, lecithin, among the Pu Luonike F-68 one or more, the weight percentage of tanshinone is 90～100% in the described TANSHINONES.
2. tanshinone emulsion according to claim 1 is characterized in that: described Emulsion is used for intravenous injection Emulsion.
3. tanshinone emulsion according to claim 2, it is characterized in that: contain 100 milligrams～1500 milligrams of tanshinones, oil for injection 100 grams～500 grams, glycerol for injection 5 grams～50 grams, emulsifying agent 5 grams～50 grams in every 1000ml used for intravenous injection Emulsion, all the other are water for injection.
4. tanshinone emulsion according to claim 3 is characterized in that: contain 500 milligrams of tanshinones, oil for injection 200 grams, glycerol for injection 22 grams, emulsifying agent 15 grams in every 1000ml used for intravenous injection Emulsion, all the other are water for injection.
5. method for preparing the used for intravenous injection tanshinone emulsion, it is characterized in that: the preparation method of described used for intravenous injection Emulsion comprises the steps:
A, take by weighing each raw material: 100 milligrams～1500 milligrams of tanshinones, oil for injection 100 grams～500 grams, glycerol for injection 5 grams～50 grams, emulsifying agent 5 grams～50 grams, all the other are water for injection, and used emulsifying agent is a fabaceous lecithin;
B, get in the oil for injection that tanshinone is dissolved in preheating, take by weighing fabaceous lecithin and glycerol again and be dissolved in an amount of water for injection that is preheated to 80 ℃;
C, above-mentioned water is changed in the tissue mashing machine, under 10000 rev/mins stirring condition, slowly drip oil phase and go into water, change stirring 10 minutes with per minute 10000, be dispersed in aqueous phase until the oil phase that is dissolved with TANSHINONES and obtain milky or orange-yellow colostrum;
D, get above-mentioned colostrum and add water for injection and make and reach full dose 1000ml, be transferred in the high pressure dispersing emulsification machine 15000Psi homogenize 3 times, the sampling and measuring particle diameter reaches below 0.5 micron to mean diameter, gets above-mentioned Emulsion embedding in infusion bottle or ampoule, inflated with nitrogen was sterilized 30 minutes down for 115 ℃, promptly.
6. anhydrous lactitol fluid composition, it is to contain right to require 1～4 each described tanshinone emulsion be main component, adds the solid preparation that acceptable accessories or complementary composition are prepared from.
7. anhydrous lactitol fluid composition according to claim 6 is characterized in that: described adjuvant or complementary composition are that cryoprotective agent, skeleton form one or more in agent, hydrophilic emulsifying agent, the acetylation monoglyceride.
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| JP2011500557A (en) * | 2007-10-11 | 2011-01-06 | マゼンス インコーポレイテッド | Pharmaceutical composition containing atomized particles of naphthoquinone compound |
| CN101422454B (en) * | 2007-10-15 | 2011-06-15 | 浙江九旭药业有限公司 | Omega-3 polyunsaturated fatty acid tanshinone IIA sub-microemulsion and preparation method thereof |
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| CN1092646A (en) * | 1992-12-24 | 1994-09-28 | 施瓦茨制药有限公司 | The lyophilizing emulsion that contains effective composition |
| CN1426782A (en) * | 2001-12-17 | 2003-07-02 | 中山大学 | Tanshinone 11A used in preparing medicine for preventing and treating atherosclerosis |
| CN1480136A (en) * | 2003-07-18 | 2004-03-10 | 中国药科大学 | Nano granules of solid lipid of tanshinone and its preparation method |
| CN1483398A (en) * | 2002-09-20 | 2004-03-24 | 西安力邦医药科技有限责任公司 | Vegetable oil fatty milk oral or injection preparation |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1092646A (en) * | 1992-12-24 | 1994-09-28 | 施瓦茨制药有限公司 | The lyophilizing emulsion that contains effective composition |
| CN1426782A (en) * | 2001-12-17 | 2003-07-02 | 中山大学 | Tanshinone 11A used in preparing medicine for preventing and treating atherosclerosis |
| CN1483398A (en) * | 2002-09-20 | 2004-03-24 | 西安力邦医药科技有限责任公司 | Vegetable oil fatty milk oral or injection preparation |
| CN1480136A (en) * | 2003-07-18 | 2004-03-10 | 中国药科大学 | Nano granules of solid lipid of tanshinone and its preparation method |
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