CN1631444A - Ultrasound contrast medium composition with phospholipid as membrane material and its preparation method - Google Patents

Ultrasound contrast medium composition with phospholipid as membrane material and its preparation method Download PDF

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CN1631444A
CN1631444A CN 200310122421 CN200310122421A CN1631444A CN 1631444 A CN1631444 A CN 1631444A CN 200310122421 CN200310122421 CN 200310122421 CN 200310122421 A CN200310122421 A CN 200310122421A CN 1631444 A CN1631444 A CN 1631444A
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composition
freeze
phospholipid
ultrasound
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CN1321697C (en
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梅兴国
李思成
赵应征
唐杰
张彦
栾新慧
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中国人民解放军军事医学科学院毒物药物研究所
成都一平医药科技发展有限公司
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Abstract

The invention relates to an ultrasound contrast medium composition with phospholipid as membrane material and its preparation method, wherein the contrast medium comprises film forming material and fluorine-carbon type inert gases, the film forming material comprises phosphatide constituent 1-10%, foaming agent 5-15%, stabilizing agent 0.5-10%, and polymeric compound 70-90%.

Description

一种以磷脂类成分为成膜材料的超声造影剂组合物及其制备方法 Ultrasound contrast agent composition and to a method of preparing phospholipid film-forming material component

技术领域 FIELD

:本发明涉及一种超声造影剂组合物,具体涉及一种以磷脂类成分为成膜材料的包裹氟碳类惰性气体的超声造影剂组合物及其制备方法。 : The present invention relates to an ultrasound contrast agent composition, particularly relates to an ultrasound contrast agent composition and a method of preparing phospholipid component is to package an inert gas forming a fluorocarbon material.

技术背景:新型超声造影剂结合超声新技术能有效增强心肌、肝、肾、脑等实质性器官的二维超声影像和血流多普勒信号,反映正常组织和病变组织(肿瘤、缺血心肌)不同的血流灌注,明显提高超声诊断的敏感性和特异性。 Background: The new ultrasound contrast agent in conjunction with the new ultrasound technology can effectively enhance the two-dimensional ultrasound imaging and Doppler blood flow signal substantive organs heart, liver, kidney, brain, reflecting the normal tissue and diseased tissue (tumor, myocardial ischemia) different blood perfusion, significantly improve the sensitivity and specificity of ultrasound diagnosis. 除此之外携带基因、药物的超声造影剂在治疗方面也有广泛的应用前景。 In addition to carrying the gene, the drug in the treatment of ultrasound contrast agents also have broad application prospects. 理想的新型超声造影剂具备以下特点:高散射性、低弥散性、低溶解性、无生物学活性(对人体无害人可自由通过毛细血管,微泡大小均匀,组织显影好,有效增强组织显影足够满足检查时间。 Ideal new ultrasound contrast agent has the following characteristics: high scattering property, low dispersibility and low solubility, biologically inactive (harmless person may freely through the capillary, uniform size microbubbles, tissue development, tissue effective to enhance The developing time sufficient for examination.

新一代超声造影剂多以含氟气体为微泡的核心,因含氟碳气体为惰性气体,分子量大,在血液中的溶解度和弥散性差,稳定性好。 Ultrasound contrast agents in the new generation of multi-core fluorine gas microbubbles, due to fluorocarbon-containing gas is an inert gas, molecular weight, solubility and dispersibility in blood poor stability. 按包裹氟碳气体的材料不同而有白蛋白类、表面活性剂类、磷脂类、高分子聚合物类。 Press wrapping material to the fluorocarbon gas while various albumins, surfactants, phospholipids, high molecular polymers. 就其在超声造影的增强显像上比较,以磷脂类成分为成膜材料造影剂有更多的优势,原因在于:(1)靶向性。 Its comparison on an enhanced ultrasound contrast imaging of the phospholipid film-forming material component based contrast agents have more advantages, because: (1) targeting. 磷脂类造影剂进入人体后,易优先被富含网状内皮细胞的组织如肝、脾及骨髓所摄取。 Phospholipids contrast agent into the body, easy to reticuloendothelial cells preferentially enriched tissues such as the liver, spleen and bone marrow ingested. (2)稳定性好。 (2) good stability. 磷脂类造影剂化学性质稳定,常温下可保存数月不变化,易于商品化;(3)安全。 Phospholipids contrast agent is chemically stable, can be stored for several months at room temperature does not change, easy commercialization; (3) safety. 构成磷脂类造影剂的磷脂膜可生物降解,对人体无害;而白蛋白类造影剂由于以人血白蛋白为载体,尚存有传播血液性疾病的危险。 Phospholipid membrane phospholipids constituting the contrast agents are biodegradable, harmless to human body; and albumin-based contrast agents due to human serum albumin as the carrier, the remaining risk of the spread of blood diseases.

发明内容 SUMMARY

:本发明的目的是提供一种改进的以磷脂类成分为成膜材料的造影剂组合物;本发明还提供一种优化的造影剂的制备方法;通过实现本发明的目的能够提高微气泡的产率,延长造影剂在组织显影的有效增强时间。 : Object of the present invention to provide an improved contrast agent composition phospholipid component of the film-forming material; the present invention also provides a method for preparing an optimized contrast agent; object achieved by the present invention can improve micro-bubble yield, prolonged time effective to enhance the contrast agent in the developing tissue.

为实现上述目的之一,本发明采用了以下的技术方案:本发明提供的以磷脂类成分为成膜材料的造影剂组合物是这样一种组合物,它使用磷脂类物质为主要成分制成一种成膜材料,将该成膜材料和氟碳类惰性气体混合一起施用达到造影的目的。 In order to achieve one of the above-described object, the present invention adopts the following technical scheme: the present invention provides phospholipid-based component is a film-forming material contrast agent composition is a composition, which made use of a phospholipid as a main component substances a film-forming material, the film-forming material and fluorocarbon inert gas mixture administered with the purpose of angiography. 在成膜材料的组成中,包括磷脂类成分、起泡剂、聚合物成份、稳定剂等成分;这些成份在造影剂组合物中所占的重量百分比为,磷脂类物质的比例为1~10重量%,起泡剂的比例为5~15重量%,稳定剂比例为0.5~10重量%,高分子聚合物构成比为70~90重量%,其余为氟碳类惰性气体。 In the composition of the film-forming material, the component comprising phospholipids, foaming agents, polymer components, stabilizers components; these components share a contrast agent composition in weight percent, the ratio of phospholipid material is 1 to 10 wt%, the proportion of foaming agent is 5 to 15% by weight, the proportion of stabilizing agent is 0.5 to 10 wt%, the constituent ratio of polymer 70 to 90% by weight, the remainder being an inert fluorocarbon gas.

以上造影剂组合物可以以单位剂量形式存在,如分装成小瓶,使用时根据病人的需要施以不同的剂量,每一剂量单位中可以含有氟碳类惰性气体0.15~0.5ml。 The above contrast agent compositions may be presented in unit dosage form, such as distributed into vials, subject to different doses according to the patient during use, each dosage unit may contain an inert gas such as a fluorocarbon 0.15 ~ 0.5ml.

所说的磷脂类成分选自至少一种以下物质:卵磷脂(PC)、氢化大豆磷脂(hydrogenated soy phosphatidylcoline,HSPC)、氢化蛋黄磷脂(hydrogenatedegg phosphatidylcoline,HEPC)、二棕榈酰磷脂酰乙醇(DPPE)、二棕榈酰磷脂酰胆碱(DPPC)、二油酰磷酯酰乙醇胺(DOPE)、聚乙二醇-二硬脂酰磷脂酰乙醇胺(PEG-DSPE)、1,2-二棕榈酰基-sn-甘油基-3-磷脂酸甘油基-钠盐(DPPG)、1,2-二硬脂酰基-sn-甘油基-3-磷脂酰胆碱(DSPC)、1,2-二棕榈酰基-sn-甘油基-3-磷脂酸-钠盐(DPPA)、1,2-二棕榈酰基-sn-甘油基-3-磷脂酰胆碱(DPPC)。 Said phospholipid component is selected from at least one of the following: phosphatidylcholine (PC), hydrogenated soy phosphatidylcholine (hydrogenated soy phosphatidylcoline, HSPC), hydrogenated egg yolk phospholipid (hydrogenatedegg phosphatidylcoline, HEPC), dipalmitoyl phosphatidyl ethanol (of DPPE) , dipalmitoyl phosphatidyl choline (of DPPC), dioleoyl phosphatidylethanolamine (DOPE), polyethylene glycol - distearoyl phosphatidyl ethanolamine (PEG-DSPE), 1,2- dipalmitoyl -sn - glycero-glycero-3-phosphatidic acid - sodium salt (DPPG), 1,2- distearoyl -sn- glycero-3-phosphatidylcholine (DSPC), 1,2- dipalmitoyl -sn - glycero-3-phosphatidic acid - sodium salt (DPPA), 1,2- dipalmitoyl -sn- glycero-3-phosphatidylcholine (DPPC).

所说的起泡剂选自至少一种非离子表面活性剂,非离子表面活性剂具体的如吐温类物质、司盘类物质;使用起泡剂的目的是为了增加在制备造影剂微泡时微泡的产出率,同时对脂膜起到稳定的作用。 Said foaming agent is selected from at least one non-ionic surfactant, a nonionic surfactant such as Tween specific substances, Span species; frothers purpose is to increase the contrast agent microbubbles prepared when the micro bubbles yield, while stabilizing effect on the lipid membrane.

所说的聚合物选自至少一种高分子聚合物,高分子聚合物具体的如泊洛沙姆(Poloxamer),高分子聚合物的使用是为脂质成份及起泡剂在构成造影剂膜时提供一个支持结构,同时对膜的稳定性也有贡献。 Said polymer is selected from at least one polymer, such as specific poloxamers (Poloxamer), a high molecular polymer is a polymer composition and foaming agent lipids constituting the membranes of contrast agents providing a support structure, while also contributing to the stability of the film.

使用的稳定剂选自聚乙二醇、单硬脂酸甘油酯、棕榈酸,优选的是聚乙二醇1500(PEG1500),应用稳定剂的目的是为了减少造影微泡的相互融合倾向,改善脂质双分子层的亲水亲脂作用,增强造影微泡的稳定性。 Stabilizer used is selected from polyethylene glycol, glyceryl monostearate, palmitic acid, preferably polyethylene glycol 1500 (PEG1500), Objective stabilizers are integrated with each other to reduce the tendency of the microbubble contrast, improved hydrophilic-lipophilic lipid bilayer, enhance the stability of contrast microbubbles.

氟碳类惰性气体在造影剂中的作用是与成膜材料共同为超声波提供一个声阻抗差较大的反射界面,所述的氟碳类惰性气体选自氟化物气体,氟化物气体具体地讲包括有全氟化碳气体、氟化硫气体等,在应用中主要为全氟丙烷、六氟化硫。 Effect of the inert gas in the fluorocarbon contrast agents with common film-forming material to provide an ultrasonic acoustic impedance difference is large reflective interface, the fluorocarbon-based gas is an inert gas selected from a fluoride, fluorine compound gas particular comprises a perfluorocarbon gas, sulfur fluoride gas, mainly in the application perfluoropropane, sulfur hexafluoride.

本发明的造影剂组合物可以采用以下的方法制备:步骤(a):将所述的成膜材料磷脂类成分、起泡剂、聚合物成份、稳定剂等成分(具体如HEPC、Poloxamer 188、Tween 80及PEG 1500)与少量无水或含微量水的非水溶剂接触,利用超声和加热方式使其成为一个均匀的溶液体系,使用的温度为45~65℃,时间为20~40分钟。 Contrast agent compositions of the invention may be prepared using the following method: Step (a): the phospholipids forming material composition, foaming agents, polymer components, stabilizers and the like of the component (such as particular HEPC, Poloxamer 188, Tween 80 and PEG 1500) in contact with a small amount of anhydrous or non-aqueous solvent containing a trace of water, and using an ultrasonic heating make it a homogeneous solution system, the temperature used is 45 ~ 65 ℃, for 20 to 40 minutes.

在上述的步骤中所述的非水溶剂为无水或含微量水的丁醇等直链或支链醇。 Described in the above step is anhydrous or non-aqueous solvent containing a trace amount of water-chain or branched butanol, straight-chain alcohols. 更优选的实施方案中所述的非水溶剂为叔丁醇。 A more preferred embodiment the non-aqueous solvent is tert-butanol. 在上述做成膜材料散布及溶解于叔丁醇等直链或支链醇时所要求的温度为45~65℃,时间为20~40分钟。 In the above made film material was dissolved in tert-butanol and dispersed like a straight-chain or branched alcohol desired temperature is 45 ~ 65 ℃, for 20 to 40 minutes.

步骤(b):将(a)溶液缓慢降温至0-4℃,使液体凝固,或先迅速放于0-4℃冷藏,然后使用超声(室温15-22℃)至乳浊液状或反复冻融方式使析出的颗粒分散均匀。 Step (b): The (a) was slowly cooled to 0-4 deg.] C, the liquid solidifies rapidly in a refrigerator before or 0-4 deg.] C, and then using ultrasound (15-22 deg.] C room temperature) or a liquid emulsion to repeated freeze way into the precipitated particles are dispersed uniformly. 超声方式可选用水浴式或探针式超声,更优选的实施方案中所述的超声方式为探针式超声。 Ultrasonically optional ultrasonic water bath or probe, a more preferred embodiment of the ultrasonic probe is ultrasonically.

步骤(c):用冷冻干燥机冷冻干燥处理,进行冷冻干燥的时间为20~25小时,冷冻干燥中所述负压吸引压力为50~120×10-3mBar,冷冻干燥温度控制为-40~-50℃。 Step (c): a drying process in a freeze dryer frozen, freeze-dried for 20 to 25 hours, freeze-dried in the suction pressure is 50 ~ 120 × 10-3mBar, freeze-drying temperature is controlled to -40 ~ -50 ℃.

步骤(d)在严格控制的无菌室内将冷冻干燥好的冻干品粉碎,分装于小瓶中制成粉针剂,向瓶内注入氟碳类惰性气体,封盖即成。 Step (d) in a sterile room strictly controlled lyophilized lyophilized good pulverization, dispensed in vials made of powder, injection fluorocarbon inert gas bottle, the closure Serve.

本发明的优越性表现在:1.微泡产出率高,微泡浓度达1×109/ml,而且微泡均一性好,直径2~6um的微泡占50~70%(见图1);2.造影剂在组织显影的有效增强时间长;3.成本低,产品造价远低于同类产品。 Advantages of the invention in the form: a high yield of microbubbles, the microbubble concentration of 1 × 109 / ml, microbubbles and good uniformity of diameter of 2 ~ 6um microbubbles from 50 to 70% (see Figure 1 ); 2. contrast agent effective to enhance the development of a long time to organize; 3. low cost, less expensive than similar products in the product.

附图说明 BRIEF DESCRIPTION

:图1为配置后不同时间的微泡形态(造影剂稀释4倍后400倍光镜下观察)。 : FIG. 1 is a form of microbubbles at different times (400 times optical microscope observation of the contrast agent was diluted 4-fold) after the configuration. a配置300min后;b配置5min后;c配置10min后;d配置20min后;e配置30min后;f配置50min后;g配置90min后;h配置120min后。 After a configuration 300min; b disposed after 5min; c arranged after 10min; d arranged after 20min; e arranged after 30min; f after 50min configuration; G arranged after 90min; 120min h after configuration.

图2为造影剂经兔耳缘静脉团注给药后对肾脏声学造影增强显像图像;图中显示的是以0.1mLl/kg的剂量分别在推注前(a)、造影剂推注后30秒(b)、1分钟(c)的肾脏显影情况。 Figure 2 is a contrast agent via rabbit ear after intravenous bolus injection administration enhanced imaging contrast echocardiography images on the kidneys; Shown is 0.1mLl / kg bolus dose, respectively before (a), after the contrast bolus 30 seconds (b), 1 minutes (c) where the developing kidney.

图3为造影剂经兔耳缘静脉团注给药后对肝脏声学造影增强显像图像,图中显示的是以0.1mL/kg的剂量分别在推注后10秒(a)、30秒(b)、1分钟(c)肝实质显影情况。 Figure 3 is a contrast agent via rabbit ear after intravenous bolus injection administration enhanced liver imaging contrast echocardiography images, is shown in FIG. 10 seconds 0.1mL / kg bolus dose, respectively (a), 30 seconds ( b), 1 minutes (c) where the developing liver.

具体实施方式 Detailed ways

:实施例1:以磷脂类成分为成膜材料的造影剂的制备称取十六酸1.5mg、单硬脂酸甘油酯2mg、聚乙二醇1500(PEG1500)120mg和泊洛沙姆188(Poloxamer 188)120mg分散于3ml的叔丁醇,水浴60℃溶解。 : Example 1: Preparation of film-forming component to phospholipid contrast agent material weighed hexadecanoic acid 1.5mg, glyceryl monostearate 2mg, polyethylene glycol 1500 (PEG1500) 120mg and poloxamer 188 (Poloxamer 188) 120mg dispersed in 3ml of t-butanol, dissolved in a water bath 60 ℃. 另称取蛋黄卵磷脂(PC)20mg分散于1ml叔丁醇中,水浴60℃超声溶解,与上液混匀,迅速于0~4℃冷置30分钟以上,液体呈凝固状,冻干20小时。 Another weighed egg yolk lecithin (PC) 20mg 1ml dispersed in tert-butanol, 60 ℃ water bath ultrasonic dissolution, mixing with the liquid, rapidly cooled to 0 ~ 4 ℃ set for 30 minutes, the liquid form of solid, lyophilized 20 hour. 称取200mg冻干样品装于5ml西林瓶中,充全氟丙烷气体,密塞,即制备得到固态的超声造影剂。 Weigh 200mg lyophilized sample mounted on a vial 5ml, perfluoropropane gas charge, Mesa, i.e. the preparation of an ultrasound contrast agent to give a solid.

使用时隔塞注入2ml生理盐水,轻轻摇动形成微泡混悬液,取样观察微泡浓度为2×107/ml,粒径分布在2~10um,泡壁厚为300-1000纳米。 2ml of physiological saline injection after a lapse of a plug used, microbubble suspension formed was gently shaken, the sample was observed microbubble concentration 2 × 107 / ml, particle size distribution of 2 ~ 10um, bubble wall thickness of 300-1000 nm.

以兔为模拟动物,采用静脉团注注射方式进行体内的超声显影效果测定,结果表明本品的微泡与市售的超声造影剂Levovist有相似的显影效果,说明本品具有优良的体内超声显影作用。 Animal simulation as rabbit using intravenous bolus effect in vivo ultrasonic determination of the developing injecting injection, and the results showed that microbubbles ultrasound contrast agents commercially available product of the present Levovist similar effect development, the product described has good intracorporeal ultrasound developing effect.

实施例2:吐温的影响称取十六酸1.5mg、单硬脂酸甘油酯2mg、聚乙二醇1500(PEG1500)120mg和泊洛沙姆188(Poloxamer 188)120mg分散于3ml的叔丁醇,水浴60℃溶解。 Example 2: Effect of Tween weighed hexadecanoic acid 1.5mg, glyceryl monostearate 2mg, polyethylene glycol 1500 (PEG1500) 120mg and poloxamer 188 (Poloxamer 188) 120mg dispersed in 3ml of t-butanol , 60 ℃ water bath to dissolve. 另称取蛋黄卵磷脂(PC)20mg和不同量的吐温80(Tween 80)分散于1ml叔丁醇中,水浴60℃超声溶解,与上液混匀,迅速于0~4℃冷置30分钟以上,液体呈凝固状,冻干20小时。 Another 20mg and said different amount of egg yolk lecithin (PC), Tween 80 (Tween 80) was dispersed in 1ml tert-butanol, 60 ℃ water bath ultrasonic dissolution, mixing with the liquid, rapidly cooled to 0 ~ 4 ℃ counter 30 min or more, the liquid form of solid, lyophilized for 20 hours. 称取200mg冻干样品装于5ml西林瓶中,充全氟丙烷气体,密塞,即制备得到固态的超声造影剂。 Weigh 200mg lyophilized sample mounted on a vial 5ml, perfluoropropane gas charge, Mesa, i.e. the preparation of an ultrasound contrast agent to give a solid.

使用时隔塞注入2ml生理盐水,轻轻摇动形成微泡混悬液,取样观察微泡微泡的浓度、平均粒径及2~8um的微泡百分比,结果见表1。 2ml of physiological saline injection after a lapse of a plug used, microbubble suspension formed was gently shaken, the sample concentration of microbubbles was observed microbubbles, average particle diameter and percentage of microbubbles 2 ~ 8um results shown in Table 1.

表1不同含量的Tween 80对微泡浓度、粒径及粒径分布的影响Tween 80含量(重量%)项目3 6 8 10 14 18 Table 1, varying amounts of Tween 80 Tween 80 content on the microbubble concentration, particle size and particle size distribution of (wt%) Item 368101418

微泡浓度4 3 5 5 7 5(107/ml)平均粒径(um) 7 7 6 6 6 62~8um的微泡百30 25 30 35 26 20分比从表1中可以看出,不同含量的Tween 80对微泡的浓度、粒径及粒径分布的影响较为显著,Tween 80的含量在8~14重量%时,微泡的浓度、平均粒径及2~8um的微泡百分比三者综合结果最好。 Microbubble concentration 4 3 5 5 7 5 (107 / ml) average particle size (um) 7 7 6 6 6 62 ~ 8um microbubbles one hundred 302,530,352,620 dividing ratio can be seen from Table 1, varying amounts of of Tween 80 on the microbubble concentration, particle size and particle size distribution is more remarkable when the content of Tween 80 was 8 to 14% by weight, the concentration of microbubbles, average particle diameter and percentage of microbubbles of three 2 ~ 8um The best overall results. 试验发现Tween 80含量越高,冻干样品粘性也相应增加,因此更优选的实施方案中所述的Tween 80的含量不超过15重量%。 It was found that the higher the content of Tween 80, lyophilized sample corresponding increase in viscosity, and therefore the more preferred embodiment the content of Tween 80 was not more than 15 wt%. 实施例2中Tween 80的含量5~15重量%的微泡的显影效果好于实施例1。 Example 2 Tween 80 content of 5 to 15 wt%, the effect of developing better microbubbles in Example 1.

实施例3:卵磷脂氢化后的影响有研究认为以氢化卵磷脂(hydrogenated egg phosphatidylcholine,HEPC)制备的长循环脂质体比未氢化卵磷脂(non-hydrogenated eggphosphatidylcholine,EPC)制备的长循环脂质体体内清除慢。 Example 3: Effect of the hydrogenated lecithin studies demonstrated that long-circulating liposomes hydrogenated lecithin (hydrogenated egg phosphatidylcholine, HEPC) prepared than unhydrogenated lecithin (non-hydrogenated eggphosphatidylcholine, EPC) long-circulating liposomes prepared slow clearance in vivo. 我们将蛋黄卵磷脂的氢化:粉状蛋黄卵磷脂(PC)加乙醇溶解,加5%钯碳,置氢化仪上氢化处理。 We hydrogenated egg yolk lecithin: powdered egg yolk lecithin (PC) was dissolved in ethanol, 5% palladium on carbon, on the opposite hydrotreatment hydrogenation apparatus. 氢化处理结束后,过滤,滤液减压旋转除去乙醇,冷冻干燥得白色氢化卵磷脂(hydrogenated egg phosphatidylcholine,HEPC)。 After completion of the hydrogenation, filtered and filtrate was removed by rotary ethanol to give a white freeze-dried hydrogenated lecithin (hydrogenated egg phosphatidylcholine, HEPC).

以氢化卵磷脂5mg取代实施例1中的蛋黄卵磷脂(PC)20mg,Tween 80 25mg其余步骤与实施例1相同,制备超声造影剂。 5mg substituted hydrogenated lecithin in Example 1 egg yolk lecithin (PC) 20mg embodiment, Tween 80 25mg remaining steps same as in Example 1, an ultrasound contrast agent was prepared. 取样观察微泡浓度为1×108/ml,粒径分布在2~8um,泡壁厚为200-800纳米。 Sampling was observed microbubble concentrations of 1 × 108 / ml, particle size distribution of 2 ~ 8um, foam thickness of 200 to 800 nanometers.

实施例3的微泡比实施例1和实施例2有更强的显影强度和更长的显影时间,说明本品比实施例1和实施例2有更好的显影效果。 Example 3 microbubbles than that of Example 1 and Example 2 have a greater strength and longer development time of development than the product described in Example 1 and Example 2 have better developing the effect.

实施例4:不同冷冻方式的影响称取十六酸1.5mg、单硬脂酸甘油酯2mg、聚乙二醇1500(PEG1500)75mg、泊洛沙姆188(Poloxamer 188)150mg、氢化卵磷脂5mg和Tween 80 25mg分散于2ml叔丁醇中,水浴60℃超声溶解。 Example 4: Effect of different ways frozen weighed hexadecanoic acid 1.5mg, glyceryl monostearate 2mg, polyethylene glycol 1500 (PEG1500) 75mg, poloxamer 188 (Poloxamer 188) 150mg, hydrogenated lecithin 5mg Tween 80 25mg and tert-butanol was dispersed in 2ml, 60 ℃ water bath sonicate to dissolve. 采用不同的冷冻方式处理(1)-10℃迅速冷冻;(2)0-4℃迅速冷藏;(3)反复冻融方式:“0-4℃凝固-水浴15-22℃超声至乳浊液状-0~4℃凝固”;(4)15-22℃室温放置至液体凝固,0-4℃冷藏10min。 Freezing different manner (1) -10 ℃ rapidly frozen; (2) 0-4 ℃ rapidly frozen; (3) freezing and thawing mode: "0-4 ℃ Solidification - ultrasound water bath at 15-22 deg.] C to a liquid emulsion -0 ~ 4 ℃ set "; placement (4) 15-22 ℃ room temperature to the solidifying liquid, 0-4 ℃ refrigerated 10min. 冻干20小时。 Freeze-dried for 20 hours. 称取200mg冻干样品装于5ml西林瓶中,充全氟丙烷气体,密塞,即制备得到固态的超声造影剂冻干粉。 Weigh 200mg lyophilized sample mounted on a vial 5ml, perfluoropropane gas charge, Mesa, i.e. the preparation of an ultrasound contrast agent to give a solid lyophilized powder.

使用时隔塞注入2ml生理盐水,轻轻摇动形成微泡混悬液。 2ml of physiological saline injection after a lapse of a plug used, gentle shaking microbubble suspension formed. 取样观察微泡的浓度、平均粒径及2~8um的微泡百分比,结果见表2。 Concentration, average particle size and the percentage of micro-bubbles observed microbubble sample 2 ~ 8um results shown in Table 2.

表2不同冷冻方式对微泡浓度、粒径及粒径分布的影响冷冻方式(4)15~22℃室温放置项目 (1)-10℃ (2)0~4℃ (3)反复 至液体迅速冷冻 迅速冷藏 冻融方式 凝固,0~4℃冷藏10min微泡浓度1 3 6 7(108/ml)平均粒径(um) 7 7 6 62~8um的微泡百30 40 55 55分比从表2中可以看出,冷冻方式不同对微泡的浓度、粒径及粒径分布的影响显著,方式(3)(反复冻融方式:0-4℃凝固-水浴15-22℃超声至乳浊液状-0-4℃凝固)和方式(4)(15-22℃室温放置至液体凝固后0-4℃冷藏10min)所得的微泡的浓度、平均粒径和2~8um的微泡百分比结果最好。 Table 2 ways frozen microbubble concentration, freeze mode of size and size distribution (4) 15 ~ 22 ℃ room temperature for item (1) -10 ℃ (2) 0 ~ 4 ℃ (3) to the liquid rapidly repeated freezing quickly thaw frozen coagulation mode, 0 ~ 4 ℃ frozen microbubble concentration 10min 1 3 6 7 (108 / ml) average particle size (um) microbubbles 7 7 6 62 ~ 8um percentage points more than 30,405,555 from the table 2 can be seen the effect of different concentration, size and size distribution of microbubbles frozen significant manner, method (3) (repeated freeze-thaw mode: 0-4 ℃ solidification - ultrasound water bath at 15-22 deg.] C to emulsion solidification liquid -0-4 deg.] C) and means (4) (15-22 deg.] C is placed to room temperature after 0-4 ℃ refrigerated liquid coagulating 10min) the concentration of the resulting microbubbles, the average particle diameter and the percentage of micro-bubbles result 2 ~ 8um the best. 方式(3)和方式(4)的显影效果好于实施例3。 Mode (3) and means (4) developing better than Example 3.

实施例5:称取泊洛沙姆188(Poloxamer 188)150mg、氢化卵磷脂5mg和Tween 80 25mg分散于2ml叔丁醇中,水浴60℃超声溶解。 Example 5: Weigh Poloxamer 188 (Poloxamer 188) 150mg, hydrogenated lecithin and Tween 80 25mg 5mg dispersed in 2ml of tert-butanol, 60 ℃ water bath sonicate to dissolve. 18-22℃室温放置至液体凝固(40min),0-4℃冷藏10min。 18-22 deg.] C to room temperature, placed in the coagulation liquid (40min), 0-4 ℃ refrigerated 10min. 冻干20小时。 Freeze-dried for 20 hours. 称取100mg冻干样品装于5ml西林瓶中,充全氟丙烷气体,密塞,即制备得到固态的超声造影剂。 Weigh 100mg lyophilized sample mounted on a vial 5ml, perfluoropropane gas charge, Mesa, i.e. the preparation of an ultrasound contrast agent to give a solid.

使用时隔塞注入1ml生理盐水,轻轻摇动形成微泡混悬液,取样观察微泡浓度为1×109/ml,粒径分布在2~6um,泡壁厚为200-500纳米。 1ml of physiological saline using a lapse of injection plug, microbubble suspension formed was gently shaken, the sample was observed microbubble concentrations 1 × 109 / ml, particle size distribution of 2 ~ 6um, bubble wall thickness of 200-500 nm.

应用验证:兔体内肝肾的超声显影成像使用实施例5制备的造影剂对兔体内肝和肾进行声学造影,以0.03ml/Kg的剂量经兔耳静脉团注给药,肝脏、肾脏约10秒时即出现明显增强显像,30秒达到高峰,有效增强时间超过1分钟。 Application verification: rabbit liver ultrasound imaging using the developing liver and kidneys of rabbits Contrast agents for ultrasound contrast prepared in Example 5 to 0.03ml / Kg dose by intravenous bolus administration of rabbit ear, the liver, kidneys about 10 That is significantly increased when developing seconds, 30 seconds peak, effective to enhance more than 1 minute.

实施例6:称取泊洛沙姆188(Poloxamer 188)150mg、氢化卵磷脂8mg和Tween 80 20mg分散于2ml无水叔丁醇中,水浴60℃超声溶解。 Example 6: weighed Poloxamer 188 (Poloxamer 188) 150mg, hydrogenated lecithin and Tween 80 20mg 8mg dispersed in 2ml of anhydrous tert-butanol, 60 ℃ water bath sonicate to dissolve. 0-4℃冷藏至液体凝固,37-39℃超声至乳浊液状,室温放置凝固。 0-4 ℃ chilled to solidification of liquid, an emulsion liquid to ultrasound 37-39 deg.] C, room temperature for solidification. 冻干20小时。 Freeze-dried for 20 hours. 称取100mg冻干样品装于5ml西林瓶中,充全氟丙烷气体,密塞,即制备得到固态的超声造影剂。 Weigh 100mg lyophilized sample mounted on a vial 5ml, perfluoropropane gas charge, Mesa, i.e. the preparation of an ultrasound contrast agent to give a solid.

使用时隔塞注入1ml生理盐水,轻轻摇动形成微泡混悬液,取样观察微泡浓度高于1×109/ml,粒径分布在2~5um,泡壁厚为200-500纳米。 1ml of physiological saline using a lapse of injection plug, microbubble suspension formed was gently shaken, the sample was observed microbubble concentrations higher than 1 × 109 / ml, particle size distribution of 2 ~ 5um, bubble wall thickness of 200-500 nm.

应用验证:兔体内肝肾的超声显影成像使用实施例6制备的造影剂显影效果好于实施例5,肾脏显像有效增强时间超过2分钟,肝实质在10分钟时显像灰阶度仍高于未造影前。 Application verification: rabbit liver in vivo imaging using an ultrasound contrast medium of the developing effects of Preparation Example 6 is better than Example 5, the developing kidney effectively enhance than 2 minutes, at 10 minutes liver imaging gray level is still high Prior to the non-contrast.

Claims (10)

1.一种以磷脂类成分为成膜材料的超声造影剂组合物,由成膜材料和氟碳类惰性气体组成,成膜材料由磷脂类成分、起泡剂、聚合物、稳定剂组成,其特征在于:成膜材料中各组分的重量百分比,磷脂类成分为1~10%,起泡剂为5~15%,稳定剂为0.5~10%,聚合物为70~90%。 A class component is a phospholipid film-forming material ultrasound contrast agent composition, the film-forming material and fluorocarbon inert gas, forming a material composition of phospholipids, a foaming agent, a polymer, a stabilizer composition, wherein: the weight percentage of each component in the film-forming material, phospholipids content of 1 to 10%, the foaming agent is 5 to 15%, the stabilizer is 0.5 to 10%, 70 to 90% polymer.
2.权利要求1的组合物,是以单位剂量形式存在的,每一单位剂量中,含有氟碳类惰性气体的量为0.15~0.5ml。 The composition of claim 1, is present in unit dosage form, each unit dosage comprising an amount of an inert fluorocarbon gas is 0.15 ~ 0.5ml.
3.权利要求1的组合物,其中磷脂类成分选自至少一种卵磷脂、氢化大豆磷脂、氢化蛋黄磷脂、二棕榈酰磷脂酰乙醇、二棕榈酰磷脂酰胆碱、二油酰磷酯酰乙醇胺、聚乙二醇-二硬脂酰磷脂酰乙醇胺、1,2-二棕榈酰基-sn-甘油基-3-磷脂酸甘油基-钠盐、1,2-二硬脂酰基-sn-甘油基-3-磷脂酰胆碱、1,2-二棕榈酰基-sn-甘油基-3-磷脂酸-钠盐、1,2-二棕榈酰基-sn-甘油基-3-磷脂酰胆碱;其中起泡剂选自至少一种非离子表面活性剂;其中聚合物选自至少一种高分子聚合物;其中稳定剂选自聚乙二醇类物质、单硬脂酸甘油酯、棕榈酸。 The composition of hydrogenated soybean phospholipid, hydrogenated egg yolk phospholipid, dipalmitoyl phosphatidyl ethanol, dipalmitoyl phosphatidyl choline, dioleoyl phosphatidyl claim 1, wherein the phospholipid component is selected from at least one of lecithin, ethanolamine, polyethylene glycol - distearoyl phosphatidyl ethanolamine, 1,2-dipalmitoyl-glycero -sn- glycero-3-phosphatidic acid - sodium salt, 1,2-distearoyl glycerol -sn- -3-phosphatidylcholine, 1,2-dipalmitoyl -sn- glycero-3-phosphatidic acid - sodium salt, 1,2-dipalmitoyl -sn- glycero-3-phosphatidylcholine; wherein the blowing agent is selected from at least one nonionic surfactant; wherein the polymer is selected from at least one polymer; wherein the stabilizer is selected from polyethylene glycol species, glyceryl monostearate, palmitic acid.
4.权利要求3的组合物,其中非离子表面活性剂为吐温类物质和司盘类物质;高分子聚合物是泊洛沙姆;聚乙二醇类物质是聚乙二醇1500~6000。 The composition of claim 3, wherein the nonionic surfactant is Tween and Span species species; poloxamer polymers; polyethylene glycol materials are the polyethylene glycol 1500 to 6000 .
5.权利要求4的组合物,其中吐温类物质是吐温80;泊洛沙姆是泊洛沙姆188;聚乙二醇1500~6000是聚乙二醇1500。 The composition of claim 4, wherein the Tween is Tween 80 species; poloxamer is poloxamer 188; polyethylene glycol 1500 to 6000 is polyethylene glycol 1500.
6.权利要求1的组合物的制备方法,其特征在于,包括以下步骤:步骤(a)将成膜材料与少量无水或含水的有机溶剂接触,利用超声和加热方式使其成为一个均匀的溶液体系;步骤(b)将(a)溶液缓慢降温使溶液凝固,或先迅速冷藏,然后使用超声至乳浊液状或反复冻融方式使析出的颗粒分散均匀;步骤(c)冷冻干燥步骤(b)的产物;步骤(d)将步骤(c)的冻干品粉碎,分装于小瓶中制成粉针剂,向瓶内注入氟碳类惰性气体,封盖。 The method of preparing a composition according to claim 1, characterized in that it comprises the following steps: (a) a film-forming material in contact with a small amount of anhydrous or aqueous organic solvent, with ultrasound and heating to become a uniform solution system; in step (b) to (a) solidifying the solution was slowly cooled, chilled or rapidly at first, and then using ultrasound or a liquid emulsion to repeated freeze-thaw embodiment the precipitated particles are dispersed uniformly; step (c) freeze-drying step ( product of b); step (d) to step (c) freeze-dried product was pulverized, dispensed in vials made powder, an inert gas is injected fluorocarbon bottle closure.
7.根据权利要求6的制备方法,其特征在于:步骤(a)中所述非水溶剂为无水或含微量水的直链或支链醇,超声和加热的条件为,温度为45~65℃,时间为20~40分钟;步骤(b)的超声方式选自水浴式或探针式超声;步骤(c)的冷冻干燥条件是冷冻干燥的时间为20~25小时,冷冻干燥中所述负压吸引压力为50~120×10-3mBar,冷冻干燥温度控制为-40~-50℃。 7. The production method according to claim 6, wherein: step (a) in the non-aqueous solvent is anhydrous or water containing a trace of a straight-chain or branched-chain alcohols, and ultrasonic heating conditions, a temperature of 45 ~ 65 ℃, time 20 to 40 min; step (b) is selected from a water bath or ultrasonically ultrasonic probe; freeze-drying conditions in step (c) is a freeze-dried for 20 to 25 hours, freeze-dried as said suction pressure of 50 ~ 120 × 10-3mBar, freeze-drying temperature is controlled to -40 ~ -50 ℃.
8.根据权利要求7的制备方法,其特征在于:步骤(a)中所述非水溶剂为无水或含微量水的叔丁醇;步骤(b)的超声方式为探针式超声。 8. The production method according to claim 7, wherein: step (a) in the non-aqueous solvent is anhydrous or water containing a trace of t-butanol; ultrasonically step (b) of the ultrasonic probe.
9.根据权利要求8的制备方法,其特征在于:所述叔丁醇,使用前应脱水处理,每毫升叔丁醇中含水量小于0.02mL。 9. The production method according to claim 8, characterized in that: the tert-butanol, should be dehydrated prior to use, per mL of t-butanol containing less than 0.02mL.
10.权利要求6的制备方法,其特征在于,泊洛沙姆188 150mg、氢化卵磷脂8mg和吐温80 20mg分散于2ml无水叔丁醇中,水浴60℃超声溶解;0-4℃冷藏至液体凝固,37-39℃超声至乳浊液状,室温放置凝固,冻干20小时;称取100mg冻干样品装于5ml西林瓶中,充全氟丙烷气体,密塞,即制备得到固态的超声造影剂。 10. The preparation process as claimed in claim 6, wherein, Poloxamer 188 150mg, hydrogenated lecithin and Tween 80 20mg 8mg dispersed in 2ml of anhydrous tert-butanol, 60 ℃ water bath ultrasonic dissolved; refrigerated 0-4 ℃ to the solidification of liquid, the liquid emulsion to ultrasound 37-39 deg.] C, room temperature for solidification and lyophilized 20 hours; lyophilized sample weighed 100mg in 5ml loading a vial, perfluoropropane gas charge, Mesa, i.e. prepared to give a solid ultrasound contrast agents.
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