IE57272B1

IE57272B1 - Ultrasound contrast agent containing microparticles and gas micro-bubbles

Info

Publication number: IE57272B1
Application number: IE835/84A
Authority: IE
Original assignee: Schering Ag
Priority date: 1983-04-15
Filing date: 1984-04-05
Publication date: 1992-07-01
Also published as: IE840835L; AU2680584A; NO841489L; EP0122624B1; JPS59205328A; FI81264B; FI81264C; DE3473828D1; CA1239092A; DK78984A; AU566928B2; DK165171B; EP0122624A2; DK165171C; EP0122624A3; ZA842801B; FI841462A0; ATE36958T1; DK78984D0; JPH0425934B2

Abstract

1. Contrast medium containing microparticles and gas bubbles for ultrasound diagnostics, characterised in that it contains microparticles of a solid surface-active substance, optionally in combination with microparticles of a non-surface-active solid, in a liquid carrier.

Description

The invention relates to contrast agents containing microparticles and gas micro-bubbles for use in ultrasound diagnosis characterized in that they contain microparticles of a solid surfactant substance, optionally combined with microparticles of a non-surfactant substance in a liquid carrier.

The examination of organs using ultrasound (sonography) is a diagnostic method which has been well established and practised for some years. Ultrasound waves in the megahertz range (above 2 megahertz with wavelengths of between 1 and 0.2 mm) are reflected at the interfaces of various types of tissue. The resulting echoes are amplified and rendered visible.

Of particular importance is the examination of the heart by this method which is known as echocardiography (Baft, J.I. et al.: Clinical echocardiography, Futura, Mount Risco, New York 1978; Kohler, E. Rlinische Echokardiographie, Enke, Stuttgart 1979; Stefan, G. et al.: Echokardiographie, Thieme, Stuttgart-New York 1981; G. Biamino, L. Lange: Echokardiographie, Hoechst AG, 1983.).

Since fluids, including blood, produce ultrasound image contrast only when there are differences in density with respect to the surroundings, possibilities were sought of rendering the blood and its circulation visible for ultrasound examination, which is also possible through adding extremely fine gas bubbles. -2Several methods of producing and stabilising gas bubbles have been described in the literature. They can be produced, for example, before injection into the * 5 bloodstream, by vigorously shaking or stirring a liquid solution, such, for exainple, as sodium chloride solution, dye solution or previously removed blood.

Although ultrasound image contrast is achieved by these methods, they have serious disadvantages which are manifested in poor reproducibility, greatly fluctuating size of the gas bubbles and a certain risk of embolism due to a proportion of large visible bubbles. Some of these disadvantages have been eliminated by other production processes, such as, for example, by the process of U.S. Patent No. 3,640,271 in which bubbles of a reproducible size are produced by filtration or by the use of direct current electrode apparatus. Against the advantage of being able to produce gas bubbles of a reproducible size is the disadvantage of the considerable technical outlay I involved.

U.S. Patent No. 4,276,885 describes the « production of gas micro-bubbles of a specific size which are surrounded by a gelatine membrane which protects their from coalescence. The prepared bubbles can be stored only in the frozen state, for example by storing at refrigerator temperature, and they must -3be raised to body temperature again before they can be used.

U.S. Patent No. 4,265,251 describes the production and use of gas micro-bubbles with a solid saccharide covering, which bubbles may be filled with a k pressurised gas. If they are under normal pressure, they can be used as ultrasound image contrast agents; when used at an elevated internal pressure, they can be used for measuring blood pressure. Although in this case the storage of the solid gas bubbles does not present any problem, the technical outlay involved in their production gives rise to high costs as a result of the complex techniques.

The risks involved with the hitherto known contrast agents for ultrasound diagnosis are caused by two factors: the size and number both of the particles of solid material and also of the gas bubbles.

The ultrasound contrast agents prepared by the previously described methods have, in all cases, possessed only some of the following properties that are required:ί .1. Exclusion of the risk of embolism (dependent on size and number of * gas bubbles and size and number of particles of solid material). 2. Reproducibility. 3. Sufficiently long stability. -44. Ability to pass through the lungs, for example in order to obtain ultrasound image contrast of the left-hand side of the heart.

. Ability to pass through the capillaries. 6. Sterility and freedom from pyrogens. 7. Easy production at reasonable cost. 8. Easy storage.

European Patent Application with publication No.52575 describes the production of ultrasound contrast agents containing gas bubbles that are supposed to possess these necessary properties. Bowever, in order to produce them, microparticles of a solid crystalline substance, such as, for example, galactose, are suspended in a liquid carrier, and the gas, vhich is adsorbed at the particle surface and is enclosed in cavities between the particles or in intercrystalline cavities, forms the gas bubbles. The resulting suspension of gas bubbles and microparticles is injected over a period of 10 minutes. Although according to European Patent Specification 52575 the suspension prepared by the described method is capable, after injection into a peripheral vein, of appearing both on the right-hand side of the heart and also, after passing through the lungs, on the left-hand side of the heart end cf rendering visible the blood there and its circulation during ultrasound examination, it was found when checked that the contrast medium prepared by the method -5described in European Application No. 52575 and injected into a peripheral vein did not in fact produce ultrasound echoes in the left-hand side of the heart. EP-A-77752 also describes the preparation of a liquid mixture for use as a contrast agent, which in turn comprises a mixture of a surfactant or an aqueous solution of the surfactant and an aqueous viscous carrier liquid.

An object of the present invention is to provide a contrast agent for ultrasound diagnosis which is be/ing capable, after/administered intravenously, of rendering visible for ultrasound the blood and its circulation conditions not only on the right-hand side of the heart but also, after passing through the capillary bed of the lungs, on the left-hand side of the heart. In addition, it should also permit the representation of the circulation of blood through other organs, such as the myocardium, the liver, the spleen and the kidneys.

According to the invention there is provided e contrast medium containing microparticles and gas bubbles for ultrasound diagnostics, wherein the medium contains microparticles of a solid surface-active sutstance, optionally in combination with micropartic1es cf c non-surface-active solid in a liquid carrier.

The ultrasound contrast agents of the present invention possess ell the above-mentioned properties that are expected of such a contrast agent. -6Surprisingly, ve have found that, by suspending microparticles of a solid surface-active substance, optionally in conjunction vith microparticles of a solid non-surface-active substance, in a liquid carrier, an ultrasound contrast agent is obtained which, after being injected into a peripheral vein, permits reproducible ultrasound images even of blood in the arterial left-hand side of the heart. Since the left-hand side of the heart can be reached with the ultrasound contrast agent of the invention after intravenous administration, ultrasound contrasts of other organs supplied with blood from the aorta, such as the myocardium, the liver, the spleen, the kidneys, inter alia, are therefore also possible after venous IB administration. The ultrasound contrast agent of the invention is, of course, also suitable for contrasts on the right-hand .side of the heart and for all other uses as an ultrasound image contrast medium.

All substances that are physiologically tolerable in the quantities used, that is, that have a low toxicity and/or are biologically degradable and the melting point of which is higher than room temperature, are suitable as the surface-active substance for the production of microparticles. Especially suitable are lecithins, lecithin fractions and their conversion products, polyoxyethylene fatty acid esters, polyoxyethylene fatty alcohol ethers, polyoxyethylated sorbitan fatty acid esters, glycerine polyethylene -7glycol oxystearate, glycerine polyethylene glycol ricinoleate, ethoxylated soya sterols, ethoxylated castor oils and their hydrogenated derivatives, cholesterol, polyoxyethylene fatty acid stearates and polyoxyethylenepolyoxypropylene polymers having a molecular weight of from 6800 to 8975, 13300 and 16250, saccharose esters, such, for example, as sugar esters, for example saccharose dipalmitate and saccharose monolaurate or saccharose glycerides and xyloglycerides, saturated or unsaturated (C^-Cjq)fatty alcohols or (C^-Cjq)-fatty acids or their metal salts, mono-, di- and tri-glycerides, sorbitan fatty acid esters, fatty acid esters of saccharose or fatty acid esters such, for example, as butyl stearate and ascorbyl palmitate; calcium stearate, the saccharose esters of lauric acid, stearic acid and palmitic acid, and ascorbyl palmitate are preferred.

Particularly suitable surfactants are those which are relatively difficultly soluble in the carrier liquid and which are relatively easily soluble in the blood. Through this jump in the dissolving behaviour of the surfactants, it is possible to control the dissolving of the solid material, which is rich in enclosed air. -8The solid surface-active substance of the contrast agent is used in a concentration of from 0.01 to 5 %, preferably from 0.04 to 0.5 %, by weight of the contrast agent.

If desired, the microparticles of the surfaceactive substance can be used in conjunction with microparticles of a physiologically tolerable nonsurface-active crystalline solid. Organic and inorganic substances can be used as the crystalline solid, for example salts such, for example, as sodium chloride, sodium citrate, 6odium acetate or sodium tartrate, monosaccharides such, for example, as gi ucose, fructose or galactose; disaccharides such, for example, as saccharose, lactose or maltose; pentoses such, for example, as arabinose, xylose or ribose; or .: -9cyclodextrines such, for example, as The microparticles may be produced by recrystallising the surface-active substances and, if . desired, non-surface-active substances under sterile conditions. They are then comminuted under sterile conditions, for example, by grinding in an air-jet mill, until the desired particle size is obtained. Preferably the microparticles should have a median particle size of less than 10 pm, advantageously less than 8 pm, more especially within the range of from 1 to 3 pm. The particle size is determined in a suitable measuring apparatus. The microparticles produced comprise either the comminuted surface-active substance alone or a mixture of the microparticles of the surfaceactive substance and a solid non-surface-active substance. In the latter case the ratio by weight of solid surface-active substance to solid non-surfaceactive substance is preferably from 0.01 to 5:100.

Both the microparticle size achieved by the comminution process and also the size of the micro25 bubbles containing a physiologically tolerable gas contained in the contrast agent of the invention ensure safe passage through the capillary system and the -10capillary bed of the lungs and preclude the occurrence of embolism.

Some of the micro-bubbles required to produce image contrast are transported by the suspended micro5 particles, adsorbed at the surface of the microparticles and enclosed in the cavities between the microparticles or enclosed in an intercrystalline manner.

The volume of gas transported by the microparticles in the form of gas micro-bubbles is from 0.02 to 0.6 ml per gram of microparticles.

Apart from its transporting function, the carrier liquid also has the function of stabilising the suspension comprising microparticles and gas micro-bubbles, for example of preventing the sedimentation of the microparticles and the coalescing of the gas microbubbles or of delaying the dissolving process of the microparticles.

There may be used as the liquid carrier, for example, water, aqueous solutions of one or more inorganic salts such, for example, as physiological sodium chloride solution and buffer solutions, aqueous solutions of mono- or di-saccharides such, for example, as galactose, glucose or lactose, monohydric or polyhydric alcohols, in so far as they are physiologically tolerable such, for example, as ethanol, propanol, isopropyl alcohol, polyethylene 11glycol, ethylene glycol, glycerine, propylene glycol, t propylene glycol methyl ester or their agueous solutions.

Water and physiological electrolyte solutions, such, for example, as physiological sodium chloride solution, and aqueous solutions of galactose and lactose, are preferred. If solutions are used, the concentration of the dissolved substance should be from 0.1 to 30 % by weight, preferably from 0.5 to 25 1 by weight, and, more especially there may be mentioned, 0.9 % aqueous sodium chloride solution or 20 % aqueous galactose solution.

The invention also provides a process for the preparation of the contrast agent of the invention wherein microparticles of a physiologically tolerable solid surface-active substance, optionally in combination with microparticles of a physiologically tolerable non-surface-active solid, are combined with a physiologically tolerable carrier liquid and shaken until a homogeneous suspension is formed.

In order to prepare the ultrasound contrast agent in a form ready for use, the sterile liquid carrier may be added to the sterile solid surface-active substance, which is in the form of microparticles and which is optionally in conjunction with microparticles of a sterile solid non-surface-active substance, and this mixture with the carrier liquid is shaken until a homogeneous suspension has formed, which takes -12approximately from 5 to 10 seconds. Immediately after its preparation, and at the latest up to 5 minutes thereafter, the resulting suspension is injected in the form of a bolus into a peripheral vein or into a catheter which is already present, from 0.01 ml to 1 ml/kg body weight being administered.

For reasons of expediency, the components necessary for the preparation of the contrast agent of the invention such, for example, as carrier liquid and microparticles of the surface-active substance, optionally in conjunction with microparticles of the solid non-surface-active substance are stored under sterile conditions in two separate vessels (A) and (B) respectively in the quantity necessary to carry out an examination. The size of vessel (B) should1 be such that the contents of vessel (A) can be transferred to (B) by means of an injection syringe and the resulting mixture can be shaken. Thus the invention also relates the to a kit for'preparation of an ultrasound contrast medium containing microparticles and gas bubbles, comprising a) a container having a volume of from 5 to 10 ml which is provided with a closure permitting the removal of the contents under sterile conditions and which is filled with 4 ml, of the liquid carrier and b) a second container having a volume of from to 10 ml which is provided with a closure permitting the removal of the contents or the addition of a mixture of substances under sterile conditions and which is filled with microparticles of a solid surface-active -13substance, optionally in combination with micro-particles of a non-surface-active solid, having an average particle size of from <1 to 10 pm, the ratio by weight of substance-active substance to optionally present non5 surface-active solid being from 0.01 to 5:100 and the content of microparticles being from 5 to 50% by weight, preferably from 9 to 40% by weight.

The use of the contrast agent of the invention is demonstrated by an echocardiographic examination of a baboon weighing 10 kg. which will now be described. ml of carrier liquid (prepared according to Example 1 A below) are removed from a phial using an injection syringe and are added to 2 g of micro-particles (prepared according to Example 1 B below) which are in a second phial, and the mixture is shaken for approximately from 5 to 10 seconds until a homogeneous suspension has formed. 2 ml of this suspension are injected into a peripheral vein (V. jugularis. brachialis or saphena) via a three-way tap having an infusion speed of at least 1 ml\sec., preferably 2-3 ml\sec. Immediately after injecting the contrast agent, 10 ml of physiological sodium chloride solution are injected at the same speed so that the contrast agent bolus is maintained as complete as possible until the right-hand side of the heart is reacted. Before, during and after injection, a commercially available transducer for echocardiography is held against the thorax of the experimental animal so that a typical cross-section is obtained through the right-hand side and the left-hand side of the heart. -14This test procedure is understood end well known to e person skilled in the ert.

If the ultrasound contrast agent reaches the righthand side of the heart, it is possible to follow in a 25 d echo image or an «-mode echo image how the blood marked by the contrast agent first reaches the level of the right-hand atrium and then the level of the righthand ventricle and the pulmonary artery, homogeneous filling occurring for approximately 10 seconds. While the cavities in the right-hand side of the heart in the ultrasound image empty again, the blood which is rendered visible with contrast agent, after passing through the lungs, appears again in the pulmonary veins and fills the left atrium, the left ventricle and the aorta homogeneously, the contrast image lasting from 2 tc 3 times longer than on the right-hand side of the heart. In addition to the representation of the blood flow through the cavities of the left-hand side of the heart, there is also a representation of the myocardium showing the circulation of the blood.

The use of the ultrasound contrast agent pf the invention is, however, not limited to rendering visible the circulation of blood in the arterial part of the heart after venous administration but is also used with outstanding success as a contrast agent for examining the right-hand side of the heart and other organs. -15The following Examples illustrate the invention, the parts and percentages being by weight unless otherwise indicated.

Example 1 A) Preparation of the carrier liquid g of galactose are dissolved in water for injection purposes, made up to a volume of 400 ml and forced through a 0.2 pin filter; 5 ml phials are each filled with 4 ml of this filtrate and sterilised for 20 minutes at 120°C.

B) Preparation of the microparticles: Under sterile conditions, 198 g of galactose are mixed intensively with 2 g of magnesium stearate by homoeopathic trituration, passed through a 0.8 mm sieve, mixed loosely and ground in an air-jet mill until the following distribution of the particle size is obtained: Median: 1.9 pm 99 % < 6 pm 90 % <3 pm.

The particle size and the distribution thereof are determined in a particle-measuring apparatus after suspension in anhydrous isopropanol. 5 ml phials are each filled with 2 g of the microparticles. -16C) For the preparation of 5 ml of the ultrasound contrast agent in a form ready for use, the contents of a phial containing carrier fluid (20 1 galactose solution in water. A) are introduced by means of ah injection syringe into a phial containing microparticles (B) and shaken until a homogeneous suspension is formed (from 5 to 10 seconds).

Example 2 A) Preparation of the carrier liquid: Water for injection purposes is used and 5 ml phials are each filled with 4 ml of the water and sterilised for 20 minutes at 120°C.

B) Preparation of the microparticles: Under sterile conditions, 198 g of galactose are mixed intensively with 2 g of ascorbyl palmitate by homoeopathic trituration and further processed as described in Example 1 under B), the following distribution of the particle size being obtained: Median: 1.9 gm 100 % pm 90 % 4. 3 pm.

The particle size is determined as described in Example 1 under B). ml phials are each filled with 1200 ma of the microparticles. -17C) For the preparation of 4.5 ml of the ultrasound contrast agent in a form ready for use, the contents of a phial containing carrier liquid (water. A) are introduced by means of an injection syringe into a phial containing microparticles (B) and shaken until a homogeneous suspension is formed (from 5 to 10 seconds).

Example 3 A) Preparation of the carrier liquid; 4.5 g of sodium chloride are dissolved in water to a volume of 500 ml and the solution is forced through a 0.2 pm filter; 5 ml phials are each filled with 4 ml of this solution and sterilised for 20 minutes at 120°C.

B) Preparation of the microparticles: Under sterile conditions, 198 g of anhydrous lactose ( C 0.3 mm) are mixed intensively with 2 g of ascorbyl palmitate by homoeopathic trituration and the mixture is further processed as described in Example 1 under E), the following distribution of the particle size being obtained: Median: 2.8 pm 100 % < 48 pm 99 % < 12 pm The particle size is determined as described in 25 Example 1 under B). -185 ml phials are each filled with 1.6 g of the ci cropar tides.

C) For the preparation of 5 ml of the ultrasound contrast agent in a form ready for use, the contents of a phial (0.9 % sodium chloride solution in water. A) are ‘introduced by means of an injection syringe into a phial containing microparticles (B) and shaken until a homogeneous suspension is formed (from 5 to 10 seconds).

Example 4 A, Preparation of the carrier liquid: In the same manner as described in Example 3 under A), 0.9 % aqueous sodium chloride solution is prepared, introduced in 4 ml portions into 5 ml phials and sterilised for 20 minutes at 120°c.

E) Preparation of the microparticles: Under sterile conditions, 199 g of e-cyclodextrine ere mixed intensively with 1 g of ascorbyl palmitate by homoeopathic trituration and the mixture is further processed as described in Example 1 under B), micro20 particles having the following size distribution being obtained: Median: 2 pm. 99 * < 6 pm 90 « <4 pm -19The particle size is determined as described in Example 1 under B). ml phials are each filled with 400 mg of the microparticles.

C) For the preparation of 4 ml of the ultrasound contrast agent in a form ready for use, the contents of a phial (0.9 % agueous sodium chloride solution, A) are introduced by means of ah injection syringe into a phial containing microparticles Example 5 A) Preparation of the carrier liquid: g of lactose are dissolved in water for injection purposes, made up to a volume of 500 ml and forced through a 0.2 pm filter; 5 ml phials are each filled with 4 ml of the solution and sterilised for 20 minutes at 120°C.

E) Preparation of the microparticles: Ascorbyl palmitate is dissolved in methanol, filtered under sterile conditions through a 0.2 pm filter, recrystallised under sterile conditions, dried and passed through a 0.8 mm sieve. The sterile ascorbyl palmitate is then ground under sterile -20conditions in an air-jet mill until the following size distribution of the particles is obtained: Median value: 1.9 pm % < 6 pm 90 % C 3 pm.

The particle size and the distribution thereof are determined in a particle-measuring apparatus after suspension in cold aqueous 0.1 % Pluronic F68 solution.

Sterile 5 ml phials are each filled with 40 mg of the microparticles.

C) For the preparation of 4 ml of the ultrasound contrast agent in a form ready for use, the contents of a phial containing carrier liquid (10 % lactose solution, A) are introduced by means of an injection syringe into a phial containing the microparticles and shaken until a homogeneous suspension is formed.

Example 6 A) Preparation of the carrier liquid: 4.5 g of sodium chloride are dissolved in water to « volume of 500 ml and the solution is forced through a 0.2 pm filter; 5 ml phials are each filled with 4 ml of this solution and sterilised for 20 minutes at 120°C.

E) Preparation of the microparticles: Under sterile conditions, a solution, filtered -21under sterile conditions, of 0.5 g of ascorbyl palmitate in 40 g of isopropanol is absorbed on 199.5 g of sterile galactose particles, the isopropanol is removed by drying at 40° and 200 torr and comminution is carried out in an air-jet mill until the following size distribution of the particles is obtained: Median value: 1.9 pm 99 % 6 pm 90 % 3 pm. The particle size and the distribution thereof are determined in a particle-measuring apparatus, for example after suspension in isopropanol. 2 g portions of the microparticles are each packed into 5 ml phials.

C) For the preparation of 5 ml of the ultrasound contrast agent in a form ready for use, the contents of a phial containing carrier liquid (0.9 % sodium chloride solution in water, A) are introduced by means of an injection syringe into a phial containing microparticles (B) and shaken until a homogeneous suspension is formed (from 5 to 10 seconds).

Example 7 A) Preparation of the carrier liquid: 4.5 g of sodium chloride are dissolved in water and made up to a volume of 500 ml; the solution is forced through a 0.2 pm filter and 5 ml phials are each -22filled with 4 ml of this solution and sterilised for 20 minutes at 120°C.

B) Preparation of the microparticles; Under sterile conditions, 199.5 g of galactose are triturated with 0.5 g of ascorbyl palmitate, mixed intensively, passed through a 0.8 mm sieve and comminuted in an air-jet mill until the following size distribution of the particles is obtained: Median value: 1.9 pm 10 99 « pm 90 % <.3 pm.

The particle size and the distribution thereof are determined in a particle-measuring apparatus, for example after suspension in isopropanol. 2 g portions of the microparticles are each packed into 5 ml phials.

C) For the preparation of 5 ml of the ultrasound contrast agent in’a form ready for use, the contents of a phial containing carrier liquid (0.9 % sodium chloride solution in water, A) are introduced by means of an injection syringe into a phial containing microparticles (B) and shaken until a homogeneous suspension is formed (from 5 to 10 seconds). -23Example 8 A, Preparation of the carrier liquid: Water for injection purposes is used and 5 ml phials are each filled with 4 ml of the water and 5 sterilised for 20 minutes at 120°C.

B) Preparation of the microparticles: Under sterile conditions, 0.5 g of saccharose monopalmitate is triturated with 199.5 g of galactose, mixed intensively, passed through a 0.8 mm, sieve and ground in an air-jet mill until the following size Λ distribution of the particles is obtained: Median value: 1.9 pm at least 99 % pm at least 90 % C 3 pm.

The particle size and the distribution thereof are determined in a particle-measurinq apparatus, for example after suspension in isopropanol. 2 g portions of the microparticles are each packed into 5 ml phials.

C) For the preparation of 5 ml of the ultrasound contrast agent in a form ready for use, the contents of a phial containing carrier liquid (sterile water for injection purposes, A) are introduced by means of an injection syringe into a phial containing microparticles (B) and shaken until a homogeneous suspension is formed (from 5 to 10 seconds). -24Example 9 A) Preparation ot the carrier liquid: ml phials are each filled with 4 ml of water used for injection purposes and sterilised for 20 minutes at 120°C.

B) Preparation of the microparticles: Under sterile conditions, a solution, filtered under sterile conditions, of 0.5 g of saccharose monopalmitate in 40 ς of isopropanol is absorbed on 199.5 g of sterile galactose particles, the isopropanol is removed by drying at 40°C and 200 torr and grinding is carried out in an air-jet mill until the following size distribution of the particles is obtained: Median value: 1.9 pm at least 99 % <£ 6 pm at least 90 % <3 pm.

The particle size and the distribution thereof are determined in a particle-measuring apparatus after suspension in isopropanol. 2 g portions of the microparticles are each packed into 5 ml phials.

C) For the preparation of 5 ml of the ultrasound contrast agent in a form ready for use, the contents of a phial containing carrier liquid (water for injection purposes, A) are introduced by means of an -25injection syringe into a phial containing microparticles (B) and shaken until a homogeneous suspension is formed (from 5 to 10 seconds).

Example 10 A) Preparation of the carrier liquid: ml phials are each filled with 4 ml of water used for injection purposes and sterilised for 20 minutes at 120°C.

B) Preparation of the microparticles: Under sterile conditions, a solution, filtered under sterile conditions, of 0.5 g of saccharose monostearate in 40 g of isopropanol is absorbed on 199.5 g of sterile galactose particles, the isopropanol is removed by drying at 40°C and 200 torr and grinding is carried out in an air-jet mill until the following size distribution of the particle is obtained: Median value: 1.9 pm at least 99 % < 6 pm at least 90 % ¢3 pm.

The particle size and the distribution thereof are determined in a particle-measuring apparatus after suspension in isopropanol. 2 g portions of the microparticles are each packed into 5 ml phials. -26C) For the preparation of 5 ml of the ultrasound contrast agent in a form ready for use, the contents of a phial containing carrier liquid (water for injection purposes. A) are introduced by means of an injection syringe into a phial containing microparticles (B) and shaken until a homogeneous suspension is formed (from 5 to 10 seconds) .

Example 11 A) Preparation of the carrier liquid: Water for injection purposes is used and 5 ml phials are each filled with 4 ml of the water and sterilised for 20 minutes at 120°C.

E) Preparation of the'micropar tides: Under sterile conditions, 0.5 g of saccharose monostearate is triturated with 199.5 g of galactose, mixed intensively, passed through a 0.8 mm sieve and ground in an air-jet mill until the following size distribution of th’e particles is obtained: Median value: 1.9 pm at least 99 % ^6 pm at least .90 % <.3 pm.

The particle size and the distribution thereof are determined in a particle-measuring apparatus after suspension in isopropanol. 2 g portions of the microparticles are each packed into 5 ml phials. -27C) For the preparation of 5 ml of the ultrasound contrast agent in a form ready for use, the contents of a phial containing carrier liquid (sterile water for injection purposes. A) are introduced by means of an injection syringe into a phial containing microparticles (B) and shaken until a homogeneous suspension is formed (from 5 to 10 seconds).

Example 12 A) Preparation of the carrier liquid: ml phials are each filled with 4 ml of water for injection purposes and sterilised for 20 minutes at 120°C. > E) Preparation of the microparticles: Under sterile conditions, a solution, filtered under sterile conditions, of 0.5 g of saccharose * distearate in 40 g of isopropanol is absorbed on 199.5 g of sterile galactose particles, the isopropanol is removed by drying at 40°C and 200 torr and grinding is carried out in an air-jet mill until the following size distribution of the particles is obtained: Median value: 1.9 pm at least 99 % < 6 pm at least 90 % pm.

The particle size and the distribution thereof are -28determined in a particle-measuring apparatus after suspension in isopropanol. 2 g portions of the microparticles ere each packed into 5 ml phials.

Cj For the preparation of 5 ml of the ultrasound 5 contrast agent in a form ready for use, the contents of a phial containing carrier liquid (water for injection purposes, A) are introduced by means of an injection syringe into a phial containing microparticles (B) and shaken until a homogeneous suspension is formed (from 5 to 10 seconds).

Example 13 A) Preparation of the carrier liquid; Water for injection purposes is used and 5 ml phials are each filled with 4 ml of the water and 15 sterilised for 20 minutes at 120°C.

E) preparation of the microparticles: Under sterile conditions, 0.5 g of saccharose cistearate is triturated with 199.5 g of galactose, mixed intensively, passed through a 0.8 mm sieve and ground in an air-jet mill until the following size distribution of the particles is obtained: Median value: 1.9 pm. at least 99 % 6 pm at least 90 % 3 pm -29The particle size and the distribution thereof are determined in a particle-measuring apparatus after suspension in isoprppanol. 2 g portions of the microparticles are each packed into 5 ml phials.

C) For the preparation of 5 ml of the ultrasound contrast agent in a form ready for use, the contents of a phial containing carrier liquid (sterile water for injection purposes. A) are introduced by means of an injection syringe into a phial containing micro10 particles (B) and shaken until a homogeneous suspension is formed (from 5 to 10 seconds).

Claims

CLA IMS

1. Contrast medium containing microparticles and gas bubbles for ultrasound diagnostics, wherein the medium contains microparticles of a solid surfaceactive substance, optionally in combination with 5 microparticles of a non-surface-active solid in a liquid carrier.

2. A contrast medium according to claim 1, wherein the medium contains as solid surface-active substance lecithins, polyoxyethylene fatty acid esters, 10 glycerine polyethylene glycol ricinoleate* cholesterol, polyoxyethylenepolyoxypropylene polymers, saccharose esters, xyloglycerides, saturated or unsaturated (C^-Cgg)fatty alcohols, saturated or unsaturated (C^-C^gJ-fatty acids or the metal salts thereof, mono-, di- and tri15 glycerides or fatty acid esters, in the form of microparticles in a quantity of from 0.01 to 101' by weight.

3. A contrast medium according to claims 1 and 2, wherein the medium certains magnesium stearate, ascorbyl palmitate, saccharose monopalmitate, saccharose 20 monostearate or saccharose distearate as solid surfaceactive substance in the form of microparticles in a concentration of from 0.01 to 51 by weight, preferably from 0.04 to 1“ by weight.

4. A contrast medium according to claim 1, 25 wherein the medium contains as optionally present microparticles of a non-surface-active solid cyclodextrins, -31monosaccharides, disaccharides, trisaccharides , polyols or inorganic or organic salts in the form of microparticles in a concentration of from 5 to 50% by weight.

5. 5. A contrast medium according to claims 1 and 4, wherein the medi urn contains as optionally present non-surface-active solid galactose, 1 actose or cAr cyclodextrin in the form of microparticles in a concentration of from 5 to 50% by weight, preferably 10 from 9 to 40% by weight.

6. A contrast medium according to claim 1, wherein the medium contains as physiologically tolerable liquid carrier water, physiological electrolyte solution, an aqueous solution of monohydric or poly15 hydric alcohols, such as glycerine, polyethylene glycol or propylene glycol methyl ester, or an aqueous solution of a mono- or disaccharide.

7. A contrast medium according to claims 1 and 6, wherein the medium contains a physiologically toler20 able liquid carrier water, physiological saline, 10% aqueous lactose solution or 20% aqueous galactose solution.

8. A contrast medium according to claim 1 4 wherein the medium contains mi cropairti cl es of magnesium 25 stearate and galactose in a 20% aqueous galactose solution.

9. A contrast medium according to claim 1 wherein the medium contains microparticles of ascorbyl .. palmitate and galactose in water. -3210. A contrast medium according to claim 1, wherein the medium contains microparticles of ascorbyl palmitate and <*-cycl odextri n in physiological saline. 11. A contrast medium according to claim 1, 5 wherein the medium contains microparticles of ascorbyl palmitate in a 10% aqueous lactose solution. 12. A contrast medium according to claim 1, wherein the medium contains microparticles of saccharose , monopalmitate and galactose in water. .!

10. 13. A contrast medium according to claim 1, wherein the medium contains microparticles of saccharose monostearate and galactose in water.

11. 14. A contrast medium according to claim 1, wherein the medium contains microparticles of saccharose

12. 15 distearate and galactose in water. 15. A kit for the preparation of an ultrasound contrast medium containing microparticles and gas bubbles, comprising a) a container having a volume of from 5 to 20 10ml which is provided with a closure permitting the removal of the contents under sterile conditions and which is filled with 4 ml, of the liquid carrier and b) a second container having a volume of from 5 to 10 ml which is provided with a closure permitting 25 the removal of the contents or the addition of a mixture of substances under sterile conditions and which is filled with microparticles of a solid surface-active substance, optionally in combination with microparticles of a non-surface-active solid, having an average particle size of from <1 to 10 pm, the ratio by weight of surface-active substance to optionally present non-surface-active solid being from 0.01 to 5:100 and the content of microparticles being from 5 to 50% by 5 weight, preferably from 9 to 40% by weight.

13. 16. Process for the preparation of a contrast medium containing microparticles and gas bubbles for ultrasound diagnostics, wherein microparticles of a physiologically tolerable solid surface-active substance, optionally in 10 combination with microparticles of a physiologically tolerable non-surface-active solid, are combined, with a physiologically tolerable carrier liquid and shaken until a homogeneous suspension is formed.

14. 17. A contrast medium according to claim 1 substantially as 15 hereinbefore described with reference to the Examples.

15. 18. A kit according to claim 15 substantially as hereinbefore described with reference to the Examples.

16. 19. A process according to claim 16 substantially as hereinbefore described with reference to the Examples.