DE19602930A1 - Porous matrices made of low molecular weight substances for the generation of stable gas bubble suspensions, their use as ultrasound contrast agents and processes for their production - Google Patents

Abstract

The invention concerns porous matrices of low molecular substances for generating stable gas bubble suspensions, their application as an ultrasound contrast medium and the process for producing the matrices and medium.

Description

The invention relates to the object characterized in the claims that is called porous matrices for the generation of stable gas bubble suspensions Use as an ultrasound contrast agent and method for producing the Matrices and means.

Ultrasound diagnostics offer the possibility without stressful ionizing radiation as with X-ray or radionuclide examinations and relatively inexpensive in Comparison to magnetic resonance imaging physiological and pathophysiological Diagnose conditions. Ultrasonic waves are based on acoustic Properties of the fabric reflect or absorb. For imaging different acoustic properties of tissues and body fluids exploited. Due to the large density difference between body tissue or liquids on the one hand and gas bubbles on the other are gases in the form of Microbubbles are particularly suitable as contrast agents for ultrasound. Ultrasound contrast agents are therefore essentially based on gas bubbles and / or gas-containing substances researched and developed.

The simplest type of ultrasound contrast media can be done by methods such as shaking, Sonication or pumping around an aqueous suspension medium between two Syringes can be obtained. The introduced bubbles can be made by suitable Additives such as surfactants and / or viscosity-increasing substances are stabilized. Such contrast agents are e.g. B. described in EP 0 077 752. Is problematic the strong dependence of the number and size of gas bubbles on the type and duration of agitation and intensity. This makes the manufacture difficult to reproduce and the risk an embolism is uncontrollable due to large gas bubbles.

A similar type of contrast medium is described in WO 93/05819, however instead of the atmospheric gases such as air, nitrogen, carbon dioxide and noble gases, gases with a certain Q factor for the production of Bladder suspensions are suggested. As a rule, these are halogenated hydrocarbons, which are characterized by low solubilities in distinguish physiological media. Perfluorinated compounds in particular suitable as exchange gases. However, in this case too - as described above - the  Bladder suspension by pumping between two syringes using a three-way valve are introduced into the suspension medium, these agents show an inhomogeneous and poorly reproducible bubble size distribution. The risk of embolism through This increases gas bubbles that are too large.

In addition to the possibility of passing through the gas bubble suspension directly before use To produce agitation of the medium, solid supports can also be formulated from which are blistered after resuspension in a suitable diluent. A microparticle suspension is used here. Such solid supports can be microparticles, for example composed of a mixture of at least one surfactant with at least one non-surfactant Solid exist, as disclosed in EP 0 365 467.

Non-surface-active solids can be used in addition to those mentioned in EP 0 365 467 Substances can also be substances that are used as X-ray contrast media (WO 93/00930 and WO 92/21382) find use, the in the case of the latter document X-ray contrast media networked with each other via functional groups and crosslinkers are.

The microparticle suspensions mentioned are ultrasound contrast media with which Contrast effects can be achieved in the arterial system. Contrast intensity and However, duration seems to be in need of improvement.

Further examples of microparticulate ultrasound contrast media are in WO 95/21631 discloses. Here, water-insoluble wall formers become one dissolved organic solvent (toluene), then in an aqueous surfactant solution emulsified and freeze-dried. Resuspending becomes a Microparticle suspension obtained which shows ultrasound contrast in vivo. The use of certain fluorinated substances is also used for the type microparticulate ultrasound contrast medium described.

For example, EP 0 554 213 discloses microparticles based on galactose which are used instead of air SF₆ included. The contrast-enhancing effects for this remedy are, however weak.

WO 95/22994 also discloses microparticles containing fluorinated substances. The bubble size distribution is standardized and the number of bubbles significantly increased, see above  that the dose necessary for an ultrasound contrast agent application is considerable can be reduced.

WO 95/03835 describes ultrasound contrast media based on particles claims that contain defined gas mixtures. The gas mixtures consist of at least one fluorinated, gas-osmotically active component and at least a conventional gas such as nitrogen, oxygen and / or carbon dioxide. The Particles are made from proteins, dextrins, starch and starch derivatives, such as. B. Hydroxyethyl starch. As stated in the scriptures, under the substances with a high molecular weight (<500,000 Daltons) preferred, since otherwise the stabilization of the gas bubbles is insufficient. Such However, substances cannot be filtered through the kidneys and have to be filtered through the Liver are broken down. This increases the dwell time in the body. The said Hydroxyethyl starch is split into substituted oligosaccharides, which primarily renally eliminated after falling below the kidney threshold. As possible Problem is the storage of hydroxyethyl starch in the cells of the reticuloendothelial system discussed. The ethyl ether bond is the enzymatic degradation not accessible. The metabolism and elimination of the Hydroxyethylglucose, as well as its possible pharmacological effect is currently not clarified [Krech, I .; Wind, S. (1995) Krankenhauspharmazie 16 (2), 62-63]. In addition, higher molecular weight substances are hidden because of their poorer ones Solubility the risk that after resuspending the contrast medium preparation particulate portions of uncontrolled size can be applied. This results in a Embolism risk to the patient.

Fluorinated gases and air-containing particles are also described in WO 95/16467 claimed, the proportion of the fluorinated component here limited to 41% is.

WO 94/09829 describes ultrasound contrast agents containing liposomes. As gas bubbles Stabilizing surfactants become phospholipids but also other heavy in water called soluble surfactants. Such liposomal systems are produced in usually by lyophilization from freeze-dry solvents such as. B. tertiary butanol or C₂Cl₄F₂. The use of organic solvents for The production of these agents requires a lot of effort Solvent recovery and makes a careful review of the product required with regard to the residual solvent content. The as a solvent halogenated hydrocarbons used, especially fluorinated  Chlorinated hydrocarbons (CFCs) such as C₂Cl₄F₂ are also because of their ozone depleting potential to be regarded as extremely critical.

The object of the present invention was therefore to produce compounds for To provide ultrasound contrast agents that have the disadvantages of the prior art overcome, d. H. the

• - the existing pharmaceutical and pharmacological problems overcome,
• - which can be produced easily and without using organic solvents,
• - generate a reproducible size and number of bubbles,
• - in dissolved form to a high contrast intensity,
• - lead to long-lasting contrast effects,
• - have a high bubble stability and beyond that
• - Renal filterable and therefore quickly excreted.

This object is achieved by the present invention.

It has been found that preparations consisting of a porous, solid, water-soluble matrix containing a low molecular weight scaffold, a surfactant and a gas, the gas being trapped in the pores of the matrix, excellent are suitable for producing a preparation for ultrasound diagnosis.

In contrast to the microparticulate contrast media of the prior art, the contrast media according to the invention are generated from a particle-free, porous, solid structure, which is referred to below as a porous matrix. To clarify the basic structural differences, reference is made to FIGS. 1 and 2. Fig. 1 shows a scanning electron micrograph of a microparticulate preparation of the prior art (EP 0365467), Fig. 2 shows a photograph of a preparation of the invention prepared according to Example 19 at the same magnification (1 cm on the pictures corresponding to 1.1 microns in the Reality). Clearly recognizable are the uniform pores from which gas bubbles are released after the matrix is dissolved. The size and number of pores are highly reproducible. The size of the bubbles is essentially limited by the pore size. The number of bubbles that can be released from the matrix is also determined by the porosity of the matrix. The parameters mentioned (bubble size and number) can be easily controlled via various production parameters and have an important influence on the effectiveness of the contrast medium.

Also, the formation of the gas bubbles is not due to agitation of the medium before Application bound, so that the gas bubbles are released in unchanged form can. The gas bubbles released are particularly advantageous with the help of Surfactants, which may be part of the matrix, can be stabilized. Thus comes in the case of the agents according to the invention usually a stabilized one Gas bubble suspension for application.

Due to the high reproducibility of the generated bubbles, the Risk of lung embolization minimized by blisters that are too large. About that In addition, the use of low molecular weight and therefore readily soluble substances is reduced the risk of matrix formation is the risk of undissolved particulate formulation components to apply.

The porous matrices according to the invention are made up of a water-soluble one Scaffolders, which usually have a molecular weight <15000 Daltons and a quickly and well water-soluble surfactant, the surfactant content in the Matrix is 0.01 to 10% (m / m). Because the scaffolders do not must necessarily be soluble in an organic solvent, opens up a wider choice of well-tolerated scaffolders.

Possible scaffolders are:
Amino acids, polyamino acids, peptides, proteins, mono, di, tri, tetra, oligo- and polymeric saccharides, as well as their derivatives, synthetic saccharides and saccharide derivatives. Examples include L-glycine, L-alanine, L-valine, L-leucine, L-isoleucine, L-phenylalanine, L-proline, L-hydroxyproline, L-serine, L-threonine, L-tryptophan, L-asparagine , L-glutamine, L-arginine, L-histidine, glycyl-glycine, glycyl-glycyl-glycine, glucose, galactose, fructose, mannose, sorbose, sucrose, lactose, maltose, trehalose, gentiobiose, lactulose, turanose, maltotriose, melibiose, Melicitose, maltotetraose, maltopentaose, stachyose, arabinose, xylose, ribose, dulcitol, xylitol, mannitol, ribitol, inositol, sorbitol, α, β, γ-cyclodextrins, hydroxypropyl-β-cyclodextrin and other derivatives as well as polymeric saccharides with a molecular weight <15,000 Dalton, such as B. dextran 8, dextrins or synthetic saccharide polymers, such as. B. Ficoll.

X-ray contrast media and contrast media are also suitable as scaffold builders magnetic resonance imaging. Iopromide, Iotrolan, Iopamidol, iohexol, as well as Gd-DTPA (gadopentetic acid), Gd-DTPA-dimeglumin salt (Magnevist), Gd-EOB-DTPA (gadoxetic acid, disodium salt) and gadobutrol.  

According to the invention, preference is given to renally unimpeded substances which can be filtered Molecular Weight <15,000 Daltons (Silbernagl S., Despopoulos A., Taschenatlas der Physiology, p. 132), whereby saccharides with at least two sugar units, X-ray contrast media and contrast media for magnetic resonance imaging in particular are suitable. The use of substances with a molecular weight <15,000 Dalton has a clear advantage over the prior art agents because renal elimination is ensured for such substances. A burden of Through long dwell times of a contrast medium component, as well as through Metabolites are thus avoided. Because with decreasing molecular weight in the As the water solubility increases, there is also the risk of application reduced from undissolved formulation components.

Suitable surfactants are water-soluble, nonionic surfactants, those with a perfluorinated hydrocarbon building block and / or with a molecular weight <15,000 daltons are preferred. Examples include sorbitan fatty acid esters, Polyoxyethylene sorbitan fatty acid esters, polyoxyethylene sorbitan fatty acid esters, Polyoxyethylene fatty acid esters, glycerol polyoxyethylene fatty acid esters, ethoxylated Mono-, di-, triglycerides, which can, if desired, be partially hydrogenated as well ethoxylated mixtures of these, ethoxylated castor oils, ethoxylated phenols, Polyoxyethylene fatty alcohol ether, polyglycerol fatty acid ester, Sorbitan perfluoro fatty acid esters, polyoxyethylene sorbitan perfluoro fatty acid esters, Polyoxyethylene sorbitol perfluoro fatty acid esters, polyoxyethylene perfluoro fatty acid esters, Polyoxyethylene perfluorinated fatty alcohol ether, polyglycerol perfluorinated fatty acid ester, Polyoxyethylene polyoxypropylene polymers and / or fluoroalkyl poly (ethylene oxide) Alcohols such as B. Zonyle®.

In addition to the gases already established in ultrasound diagnostics, the gases such as Nitrogen, oxygen, CO₂ or air, especially fluorinated gases. Surprisingly, when using the "classic" gases, they become stronger and longer lasting in vivo contrast effect was observed than that of the particulate Preparations of the prior art can be achieved.

When using fluorinated gases, surprisingly, the use of Gas mixtures as required in the preparations of the prior art are to be waived.  

The following are preferred according to the invention, gaseous at room temperature and normal pressure:
Tetrafluorallenes, hexafluoro-1,3-butadiene, decafluorobutane, perfluoro-1-butenes, perfluoro-2-butenes, perfluoro-2-butyne, octafluorocyclobutane, perfluorocyclobutene, perfluorocyclopentane, perfluorodimethylamine, hexafluoroethane, tetrafluoromethane, pentafluoromethane, pentafluoromethane, pentafluoromethane, pentafluoromethane, Perfluoropropane and perfluoropropylene.

Among the above, the perfluorinated substances are particularly preferred.

Another aspect of the invention relates to a method for producing the matrices according to the invention. The matrices according to the invention can be produced with less effort under aseptic conditions by first providing an aqueous scaffold solution, to which gas-bubble-stabilizing surfactants are added with particular advantage. The separated or combined solutions can first be sterile filtered, then the mixture thus prepared is quickly frozen. The removal of the water takes place under conditions which allow a direct transition of the ice into the gaseous state without passing through the liquid state of matter. Possible suitable conditions can be found in the phase diagram of the water (see FIG. 3). What remains is a porous matrix that is aerated with the desired gas. For complete gas exchange (ie to remove the residual air or water vapor contained in the pores of the matrix), repeated evacuation with subsequent pressure compensation is recommended. In order to obtain a gas phase that is as pure as possible, a vacuum of <0.1 mbar should exist immediately before the pressure equalization. The production is expediently already carried out in containers which can be closed under the desired gas phase and which can later be used directly as part of a kit.

A particular advantage of the method according to the invention is that Use of organic solvents can be dispensed with. Also unnecessary a technologically complex processing, as in the case of bad or slowly soluble or only water-dispersible substances would be required. Residual solvent contents as a critical quality feature therefore play a role for the agents according to the invention do not matter. This is both from an ecological perspective, as well Also of considerable advantage in terms of product safety. So many stand organic solvents even in the smallest amounts or concentrations below that Suspected carcinogenicity and / or mutagenicity.  

The matrices of the invention can be added by adding an aqueous Medium easily produce the desired, particle-free ultrasound contrast media. The aqueous medium is added immediately before use by the treating doctor. The aqueous medium can contain the auxiliaries customary in galenics, such as B. isotonizing and viscosity-increasing additives. A shake the matrix mixed with the fluid is not required. The so produced Contrast agents are characterized by the fact that they are blood isotonic or almost blood isotonic are. They can be injected immediately after resuspending. The The concentration of the contrast media is 10 to 600 mg, preferably 50 to 400 mg Matrix material per milliliter of suspension. Depending on the application, the funds are in a dose of 0.01 to 0.20 ml / kg body weight.

The agents according to the invention are for all imaging modes of sonography, such as e.g. B. M, B, Doppler mode but also for modes in which non-linear effects are used be like B. Harmonic and Harmonic Power Mode equally suitable and can be produced with high reproducibility.

The bubble numbers generated from the matrix are significantly higher than those of the mean of the State of the art, the agents therefore show significantly improved contrast effects, in particular, the time window available for the investigation was clear can be extended (see also in-vivo experiments 41-52).

The following examples serve to explain the Subject of the invention, without wishing to restrict it to these.

example 1

20 g of dextran (MW: ∼ 1200 g / mol) are mixed with 0.2 g of Zonyl® FSO-100 (MG: ∼ 725 g / mol) and 80 g of water. The mixture is stirred until completely dissolved. The solution is filled at 5 g each and frozen with liquid nitrogen. After Removal of water under conditions that make a direct transition from the solid in the gaseous state without allowing the liquid state of matter is passed, a gas exchange with decafluorobutane is carried out. It remains a porous matrix, from which after resuspending in 10 ml of water per Grams of matrix material 12.3 × 10⁹ bubbles in the range of 0.56-7.46 microns generated can be. The number and size of bubbles was determined with a Laser diffractometer from Melvern Instruments, type Master Sizer 1000.

Example 2

10 g of dextran (MW: ∼ 1200 g / mol) are mixed with 0.1 g of Zonyl® FSO-100 (MG: ∼ 725 g / mol) and 90 g of water are added. The procedure is then as described in Example 1 method. After resuspending in 10 ml of water, per gram of substance 3.7 × 10⁹ bubbles released in the range of 0.56-7.46 µm.

Example 3

20 g of dextran 8 (MW: ∼ 8000 g / mol) are mixed with 0.2 g of Zonyl® FSO-100 (MG: ∼ 725 g / mol) and 80 g of water. The procedure is then as described in Example 1 method. After resuspending in 10 ml of water, per gram of substance 10.5 × 10⁹ bubbles in the range of 0.56-7.46 µm released.

Example 4

30 g of raffinose (MW: 594 g / mol) are mixed with 0.3 g of Zonyl® FSO-100 (MW: ∼ 725 g / mol) and 70 g of water. The procedure is then as described in Example 1 method. After resuspending in 10 ml of water, a preparation is obtained which with an osmolality of 262 mosmol is almost isotonic. Be per gram of substance 14.6 × 10⁹ bubbles in the range of 0.56-7.46 µm released.  

Example 5

20 g of trehalose (MW: 342 g / mol) are mixed with 0.2 g of Zonyl® FSO-100 (MW: ∼ 725 g / mol) and 80 g of water. The procedure is then as described in Example 1 method. After resuspending in 10 ml of water, a preparation is obtained which with an osmolality of 272 mosmol is almost isotonic. Be per gram of substance 9.4 × 10⁹ bubbles in the range of 0.56-7.46 µm released.

Example 6

20 g maltose (MG: 342 g / mol) are mixed with 0.2 g Zonyl® FSO-100 (MG: ∼ 725 g / mol) and 80 g of water are added. The procedure is then as described in Example 1 method. After resuspending in 10 ml of water, a preparation is obtained which isotonic with an osmolality of 281 mosmol. Be per gram of substance 8.2 × 10⁹ bubbles in the range of 0.56-7.46 µm released.

Example 7

40 g of maltooligosaccharide (MW: ∼ 684 g / mol) are mixed with 0.4 g of Zonyl® FSO-100 (MW: ∼ 725 g / mol) and 60 g of water are added. Then, as in Example 1 proceed as described. After resuspending in 10 ml of water, a preparation is made obtained, which is almost isotonic with an osmolality of 314 mosmol. Per gram Substance 30.0 × 10⁹ bubbles in the range of 0.56-7.46 microns are released.

Example 8

20 g of maltooligosaccharide (MW: ∼ 684 g / mol) are mixed with 0.2 g of Zonyl® FSO-100 (MW: ∼ 725 g / mol) and 80 g of water are added. It will be until full resolution touched. The solution is filled at 10 g and frozen with liquid nitrogen. After the water has been completely removed, a porous matrix remains. In the Resuspending in 10 ml of water will give a preparation containing a Osmolality of 310 mosmol is almost isotonic. 19.1 × 10⁹ per gram of substance Bubbles released in the range of 0.56-7.46 µm.  

Example 9

30 g melezitose (MW: 522 g / mol) are mixed with 0.3 g Zonyl® FSO-100 (MG: ∼ 725 g / mol) and 70 g of water. The procedure is then as described in Example 1 method. After resuspending in 10 ml of water, a preparation is obtained which with an osmolality of 315 mosmol is almost isotonic. Be per gram of substance 4.3 × 10⁹ bubbles released in the range of 0.56-7.46 µm.

Example 10

30 g melibiose (MW: 360 g / mol) are mixed with 0.3 g Zonyl® FSO-100 (MG: ∼725 g / mol) and 70 g of water are added. The mixture is stirred until completely dissolved. The solution is filled with 4 g each and frozen with liquid nitrogen. After When the water is completely removed, a porous matrix remains. In the Resuspending in 8 ml of water will result in a preparation containing a Osmolality of 306 mosmol is almost isotonic. 4.3 × 10⁹ per gram of substance Bubbles released in the range of 0.56-7.46 µm.

Example 11

30 g maltotriose (MW: 504 g / mol) are mixed with 0.3 g Zonyl® FSO-100 (MG: ∼ g / mol) and 70 g of water are added. The mixture is stirred until completely dissolved. The solution is filled at 3 g each and frozen with liquid nitrogen. After When the water is completely removed, a porous matrix remains. In the Resuspending in 6 ml of water will result in a preparation containing a Osmolality of 296 mosmol is isotonic. 3.6 × 10⁹ bubbles are formed per gram of substance released in the range of 0.56-7.46 µm.

Example 12

30 g Gd-EOB-DTPA (MW: 726 g / mol) (Gadoxetic acid, disodium) are mixed with 0.3 g Zonyl® FSO-100 (MW: ∼725 g / mol) and 70 g water. It will be up to complete dissolution stirred. The solution is filled at 3 g and filled with liquid Frozen nitrogen. After the water has been completely removed, one remains  porous matrix. When resuspending in 10 ml of water, a preparation is used obtained, which is almost isotonic with an osmolality of 344 mosmol. Per gram Substance 24.6 × 10⁹ bubbles in the range of 0.56-7.46 microns are released.

Example 13

30 g Gadobutrol (MW: 605 g / mol) are mixed with 0.3 g Zonyl® FSO-100 (MG: ∼725 g / mol) and 70 g of water are added. The mixture is stirred until completely dissolved. The solution is filled at 3 g each and frozen with liquid nitrogen. After When the water is completely removed, a porous matrix remains. In the Resuspending in 5 ml of water will give a preparation containing a Osmolality of 269 mosmol is almost isotonic. 20, 1 × 10⁹ are per gram of substance Bubbles released in the range of 0.56-7.46 µm.

Example 14

30 g gadopentetic acid (MW: 548 g / mol) are mixed with 0.3 g Zonyl® FSO-100 (MW: ∼725 g / mol) and 70 g of water were added. It will be until full resolution touched. The solution is filled at 3 g each and frozen with liquid nitrogen. After the water has been completely removed, a porous matrix remains. In the Resuspending in 7 ml of water will give a preparation containing a Osmolality of 282 mosmol is isotonic. 29, 1 × 10⁹ bubbles are formed per gram of substance released in the range of 0.56-7.46 µm.

Example 15

30 g of iopamidol (MW: 777 g / mol) are mixed with 0.3 g of Zonyl® FSO-100 (MG: ∼725 g / mol) and 70 g of water are added. The mixture is stirred until completely dissolved. The solution is filled with 4 g each and frozen with liquid nitrogen. After When the water is completely removed, a porous matrix remains. In the Resuspending in 5 ml of water will give a preparation containing a Osmolality of 268 mosmol is almost isotonic. Per gram of substance are 26.3 × 10⁹ Bubbles released in the range of 0.56-7.46 µm.  

Example 16

15 g of iopamidol (MW: 777 g / mol) are mixed with 0.15 g of Zonyl® FSO-100 (MG: ∼725 g / mol) and 85 g of water were added. The mixture is stirred until completely dissolved. The solution is filled at 8 g each and frozen with liquid nitrogen. After When the water is completely removed, a porous matrix remains. In the Resuspending in 5 ml of water will give a preparation containing a Osmolality of 343 mosmol is almost isotonic. 16.9 × 10⁹ per gram of substance Bubbles released in the range of 0.56-7.46 µm.

Example 17

30 g iohexol (MW: 821 g / mol) are mixed with 0.3 g Zonyl® FSO-100 (MG: ∼725 g / mol) and 70 g of water are added. The procedure is then as described in Example 1 method. After resuspending in 5 ml of water, a preparation is obtained which with an osmolality of 264 mosmol is almost isotonic. Be per gram of substance 24.4 × 10⁹ bubbles released in the range of 0.56-7.46 µm.

Example 18

20 g Iotrolan (MW: 1626 g / mol) are mixed with 0.2 g Zonyl® FSO-100 (MG: ∼725 g / mol) and 80g water added. The mixture is stirred until completely dissolved. The Solution is filled at 10 g and frozen with liquid nitrogen. After When the water is completely removed, a porous matrix remains. In the Resuspending in 3 ml of water will result in a preparation containing a Osmolality of 256 mosmol is almost isotonic. 10.3 × 10⁹ per gram of substance Bubbles released in the range of 0.56-7.46 µm.

Example 19

40 g of iopromide (MW: 791 g / mol) are mixed with 0.4 g of Zonyl® FSO-100 (MG: ∼725 g / mol) and 60 g of water are added. The procedure is then as described in Example 1 method. After resuspending in 6 ml of water, a preparation is obtained which isotonic with an osmolality of 289 mosmol. Be per gram of substance 22.4 × 10⁹ bubbles in the range of 0.56-7.46 µm released.  

Example 20

20 g of iopromide (MW: 791 g / mol) are mixed with 0.2 g of Zonyl® FSO-100 (MG: ∼725 g / mol) and 80 g of water are added. The mixture is stirred until completely dissolved. The solution is filled at 10 g and frozen with liquid nitrogen. After When the water is completely removed, a porous matrix remains. In the Resuspending in 6 ml of water will result in a preparation containing a Osmolality of 291 mosmol is isotonic. 21.9 × 10⁹ bubbles are formed per gram of substance released in the range of 0.56-7.46 µm.

Example 21

23 g Magnevist® (MG: 938 g / mol) are mixed with 0.23 g Zonyl® FSO-100 (MG: ∼725 g / mol) and 77 g of water were added. The mixture is stirred until completely dissolved. The solution is filled at 3 g each and frozen with liquid nitrogen. After When the water is completely removed, a porous matrix remains. In the Resuspending in 5 ml of water will give a preparation containing a Osmolality of 342 mosmol is almost isotonic. 6.16 × 10⁹ per gram of substance Bubbles released in the range of 0.56-7.46 µm.

Example 22

40 g iopromide (MW: 791 g / mol) are mixed with 0.4 g Triton®-X-100 (MG: :874 g / mol) and 60 g of water are added. The procedure is then as described in Example 1 method. After resuspending in 6 ml of water, a preparation is obtained which isotonic with an osmolality of 290 mosmol. Be per gram of substance 9.56 × 10⁹ bubbles in the range of 0.56-7.46 µm released.

Example 23

40 g of iopromide (MW: 791 g / mol) are mixed with 0.4 g of Tween®20 (MG: 718 g / mol) and 60 g of water are added. The procedure is then as described in Example 1. After resuspending in 6 ml of water, a preparation is obtained which is mixed with a  Osmolality of 289 mosmol is isotonic. 16.55 × 10⁹ per gram of substance Bubbles released in the range of 0.56-7.46 µm.

Example 24

40 g of iopromide (MW: 791 g / mol) are mixed with 0.4 g of Cremophor® RH 40 (MG: ∼ 2700 g / mol) and 60 g of water. The procedure is then as described in Example 1 method. After resuspending in 6 ml of water, a preparation is obtained which isotonic with an osmolality of 291 mosmol. Be per gram of substance 11.05 × 10⁹ bubbles in the range of 0.56-7.46 µm released.

Example 25

40 g of iopromide (MW: 791 g / mol) are mixed with 0.4 g of Rewoderm® Li 48-50 (MG: ∼ 3800 g / mol) and 60 g of water. The procedure is then as described in Example 1 method. After resuspending in 6 ml of water, a preparation is obtained which isotonic with an osmolality of 288 mosmol. Be per gram of substance 24.05 × 10⁹ bubbles in the range of 0.56-7.46 µm released.

Example 26

40 g of iopromide (MW: 791 g / mol) are mixed with 0.4 g of Solutol® HS 15 (MG: ∼ 1000 g / mol) and 60 g of water are added. The procedure is then as described in Example 1 method. After resuspending in 6 ml of water, a preparation is obtained which isotonic with an osmolality of 289 mosmol. Be per gram of substance 13.46 × 10⁹ bubbles in the range of 0.56-7.46 µm released.

Example 27

40 g iopromide (MW: 791 g / mol) are mixed with 0.4 g Lutrol® F 68 (MG: ∼ 8600 g / mol) and 60 g of water are added. The procedure is then as described in Example 1 method. After resuspending in 6 ml of water, a preparation is obtained which isotonic with an osmolality of 290 mosmol. Be per gram of substance 17.68 × 10⁹ bubbles in the range of 0.56-7.46 microns released.  

Example 28

40 g of iopromide (MW: 791 g / mol) with 0.4 g Span® 85 (MG: 1028 g / mol) and 60 g of water are added. The procedure is then as described in Example 1. After resuspending in 6 ml of water, a preparation is obtained which is mixed with a Osmolality of 291 mosmol is isotonic. 20.45 × 10⁹ per gram of substance Bubbles released in the range of 0.56-7.46 µm.

Example 29

30 g iohexol (MW: 821 g / mol) are mixed with 0.3 g Zonyl®-FSN (MG: ∼ 950 g / mol) and 70 g of water are added. The procedure is then as described in Example 1. After resuspending in 5 ml of water, a preparation is obtained which is mixed with a Osmolality of 269 mosmol is almost isotonic. Be per gram of substance 23.81 × 10⁹ bubbles in the range of 0.56-7.46 microns released.

Example 30

30 g Gadobutrol (MW: 605 g / mol) are mixed with 0.3 g Solutol® HS 15 (MG: ∼ 1000 g / mol) and 70 g of water. The mixture is stirred until completely dissolved. The solution is filled at 3 g each and frozen with liquid nitrogen. After When the water is completely removed, a porous matrix remains. In the Resuspending in 5 ml of water will give a preparation containing a Osmolality of 271 mosmol is almost isotonic. 6.64 × 10⁹ per gram of substance Bubbles released in the range of 0.56-7.46 µm.

Example 31

40 g of iopromide (MW: 791 g / mol) are mixed with 0.4 g of Zonyl® FSO-100 (MW: ∼ 725 g / mol) and 60 g of water. The procedure is then as described in Example 1 method. After drying, gas exchange with hexafluoroethane performed. A preparation is used when resuspending in 6 ml of water  obtained which is isotonic with an osmolality of 289 mosmol. Per gram of substance 4.07 × 10⁹ bubbles in the range of 0.56-7.46 µm are released.

Example 32

40 g iopromide (MW: 791 g / mol) are mixed with 0.4 g Triton®-X-100 (MG: 874 g / mol) and 60 g of water are added. The procedure is then as described in Example 1 method. After drying, gas exchange with hexafluoroethane performed. A preparation is used when resuspending in 6 ml of water obtained which is isotonic with an osmolality of 290 mosmol. Per gram of substance 3.01 × 10⁹ bubbles in the range of 0.56-7.46 microns are released.

Example 33

40 g of iopromide (MW: 791 g / mol) are mixed with 0.4 g of Tween® 20 (MG: 718 g / mol) and 60 g of water are added. The procedure is then as described in Example 1 method. After drying, gas exchange with hexafluoroethane performed. A preparation is used when resuspending in 6 ml of water obtained which is isotonic with an osmolality of 289 mosmol. Per gram of substance 4.27 × 10⁹ bubbles in the range of 0.56-7.46 µm are released.

Example 34

40 g of iopromide (MW: 791 g / mol) are mixed with 0.4 g of Cremophor® RH 40 (MG: ∼ 2700 g / mol) and 60 g of water. The procedure is then as described in Example 1 method. After drying, gas exchange with hexafluoroethane performed. A preparation is used when resuspending in 6 ml of water obtained which is isotonic with an osmolality of 291 mosmol. Per gram of substance 1.76 × 10⁹ bubbles in the range of 0.56-7.46 µm are released.

Example 35

40 g of iopromide (MW: 791 g / mol) are mixed with 0.4 g of Rewoderm® Li 48-50 (MG: ∼ 3800 g / mol) and 60 g of water. The procedure is then as described in Example 1  method. After drying, gas exchange with hexafluoroethane performed. A preparation is used when resuspending in 6 ml of water obtained which is isotonic with an osmolality of 288 mosmol. Per gram of substance 9.58 × 10⁹ bubbles in the range of 0.56-7.46 µm are released.

Example 36

40 g iopromide (MW: 791 g / mol) are mixed with 0.4 g Solutol® HS 15 (MG: ∼ 1000 g / mol) and 60 g of water are added. The procedure is then as described in Example 1 method. After drying, gas exchange with hexafluoroethane performed. A preparation is used when resuspending in 6 ml of water obtained which is isotonic with an osmolality of 289 mosmol. Per gram of substance 2.52 × 10⁹ bubbles in the range of 0.56-7.46 µm are released.

Example 37

40 g iopromide (MW: 791 g / mol) are mixed with 0.4 g Lutrol® F 68 (MG: ∼ 8600 g / mol) and 60 g of water are added. The procedure is then as described in Example 1 method. After drying, gas exchange with hexafluoroethane performed. A preparation is used when resuspending in 6 ml of water obtained which is isotonic with an osmolality of 290 mosmol. Per gram of substance 4.32 × 10⁹ bubbles in the range of 0.56-7.46 µm are released.

Example 38

40 g of iopromide (MW: 791 g / mol) are mixed with 0.4 g Span® 85 (MG: 1028 g / mol) and 60 g water added. The procedure is then as described in Example 1. After drying, a gas exchange with hexafluoroethane is carried out. When resuspending in 6 ml of water, a preparation is obtained which is mixed with a Osmolality of 291 mosmol is isotonic. 6.65 × 10⁹ bubbles are formed per gram of substance released in the range of 0.56-7.46 µm.  

Example 39

30 g iohexol (MW: 821 g / mol) are mixed with 0.3 g Zonyl®-FSN (MG: ∼ 950 g / mol) and 70 g of water are added. The procedure is then as described in Example 1. After drying, a gas exchange with hexafluoroethane is carried out. When resuspending in 5 ml of water, a preparation is obtained which is mixed with a Osmolality of 269 mosmol is almost isotonic. 8.25 × 10 Substanz per gram of substance Bubbles released in the range of 0.56-7.46 µm.

Example 40

30 g Gadobutrol (MW: 605 g / mol) are mixed with 0.3 g Solutol® HS 15 (MG: ∼ 1000 g / mol) and 70 g of water. The mixture is stirred until completely dissolved. The solution is filled at 3 g each and frozen with liquid nitrogen. The procedure is then as described in Example 1. After done Drying, gas exchange with hexafluoroethane is carried out. In the Resuspending in 5 ml of water will give a preparation containing a Osmolality of 271 mosmol is almost isotonic. Per gram of substance 2.11 × 10⁹ Bubbles released in the range of 0.56-7.46 µm.

Example 41

In vivo application of an agent according to the invention prepared according to Example 22:
A beagle dog [female, 11.2 kg body weight (hereinafter KGW)] is anesthetized (inhalation anesthesia approx. 2/3 oxygen, approx. 1/3 N₂O 1.5-2% enflurane; spontaneous breathing) and for a sonographic examination of the heart prepared. The examination is carried out using an HP ultrasound system (type 77020 E, 5 MHz transducer) in B mode. The test animal receives an intravenous application of the test substance (agent according to the invention produced according to Example 22). A contrast medium, which was produced analogously to WO 95/22994 (Example 12), serves as the reference substance.

The doses used are 0.1 ml / kg body weight both for the agent according to the invention and for the reference substance. The result is shown in the form of the intensity-time profiles in FIG. 4. The upper (slower falling) curve corresponds - as in the following FIGS. 5-8 - to the preparation according to the invention. It can be clearly seen that the agents according to the invention show a longer lasting contrast after intravenous injection than the agents of the prior art. These contrasting properties give the doctor longer periods of time for the examination (examination window). Furthermore, the need for a possible subsequent injection is significantly reduced, which reduces the burden on the patient and contributes to a favorable cost / benefit ratio.

Example 42

In vivo application of an agent according to the invention prepared according to Example 27:
The procedure is as described in Example 41. A preparation prepared as described in Example 27) serves as the test substance, and an agent from WO 95/22994 (Example 12) was used as reference. The doses for reference and test substance were identical and each was 0.1 ml / kg body weight.
The intensity-time profiles are shown in FIG. 5.

Example 43

In Vivo Use of an Agent According to the Invention Manufactured According to Example 28:
The procedure is as described in Example 41, but the test animal had a body weight of 11.9 kg. A preparation prepared as described in Example 28) serves as the test substance, and an agent from EP 0365467 (Example 1) was used. The doses used were 0.05 ml / kg body weight for the agent according to the invention according to Example 28 and 0.2 ml / kg body weight for the reference. The result of the examination is shown in the form of the intensity-time profiles in FIG. 6. In this case too, despite the low dose, a more intense and longer-lasting contrast is observed for the preparation according to the invention.

Example 44

In vivo application of an agent according to the invention prepared according to Example 29:
The procedure is as described in Example 43. A preparation prepared as described in Example 29) serves as the test substance, and an agent from EP 0365467 (Example 1) was used as a reference.

The doses used were 0.05 ml / kg body weight for the agent according to the invention and 0.2 ml / kg body weight for the reference. The result of the examination is shown in the form of the intensity-time profiles in FIG. 7.

Example 45

In vivo application of an agent according to the invention prepared according to Example 19:
A beagle dog (female, 9.7 kg body weight) is anesthetized and the procedure described in Example 41 is followed. A preparation prepared according to Example 19 serves as the test substance, and an agent from WO 95/22994 (Example 12) was used as a reference. The doses for reference and test substance were 0.1 ml / kg body weight.

The intensity-time profiles are shown in FIG. 8.

Example 46

In vivo application of an agent according to the invention prepared according to Example 5:
The procedure is as described in Example 45. A preparation prepared as described in Example 5) serves as the test substance, and an agent from WO 95/22994 (Example 12) was used as a reference. The doses for reference and test substance were identical and were 0.1 ml / kg body weight.

The result of the investigation is shown in FIG. 9. The heart of the dog was shown. In detail show:

• (a) pre-contrast
• (b) maximum contrast of the reference substance  
• (c) maximum contrast of the test substance
• (d) Contrast 1 minute after maximum contrast (reference)
• (e) Contrast 1 minute after maximum contrast (test substance)

As can be seen in the figure, the superiority of the Preparations according to the invention in particular in the case of a longer examination.

Example 47

In vivo application of an agent according to the invention prepared according to Example 11:
The procedure is as described in Example 41. A preparation prepared as described in Example 11) serves as the test substance, and an agent from WO 95/22994 (Example 12) was used as reference. The doses for reference and test substance were identical and were also 0.1 ml / kg body weight.

The result of the investigation is shown in FIG. 10. The meanings of numbers (a) to (e) correspond to those in Fig. 9

Example 48

In vivo application of an agent according to the invention produced according to one of Examples 31, 35, 39.

A beagle dog (female, 9.6 kg body weight) is anesthetized (inhalation anesthesia approx. 2/3 oxygen, approx. 1/3 N₂O, 1.5-2% enflurane, spontaneous breathing) and for one Prepared sonographic examination of the heart. The examination takes place with an ultrasound system of the brand HP (type 77020 E, 5 MHz transducer) in the B- Fashion. The test animal receives an intravenous application of one Agent according to the invention prepared according to one of Examples 31, 35, 39 and as Reference an injection of a contrast medium according to the prior art, which analog WO 95/11994 (Example 12) has been produced.

The doses used were 0.1 ml / kg body weight for the agents and for the reference. The result is in the form of the intensity values in Table 1 shown. It is also clearly evident here that the agents according to the invention are a  show significantly higher contrast levels after intravenous injection than that Reference means.

Table 1

Example 49

In vivo application of an agent according to the invention produced according to one of Examples 4, 8, 9.

A beagle dog (female, 9.7 kg body weight) is anesthetized (inhalation anesthesia approx. 2/3 oxygen, approx. 1/3 N₂O, 1.5-2% enflurane, spontaneous breathing) and for one Prepared sonographic examination of the heart. The examination takes place with an ultrasound system of the brand HP (type 77020 E, 5 MHz transducer) in the B- Fashion. The test animal receives an intravenous application of one Agent according to the invention prepared according to one of Examples 4, 8, 9 and as Reference an injection of a contrast medium according to the prior art, which analog WO 95/22994 (Example 12) has been produced.

The doses used were 0.1 ml / kg body weight for the agents and for the reference. The result is in the form of the area values under the intensity-time Curve (density units × sec) shown in Table 2. It is clearly recognizable that the  Agents according to the invention have a higher surface level after intravenous injection demonstrate.

Table 2

Example 50

In vivo application of an agent according to the invention produced according to one of the Examples 8 and 19.

A beagle dog (male, 15.5 kg body weight) is anesthetized (inhalation anesthesia 23% oxygen, 1-3% enflurane, balance nitrogen; Spontaneous breathing) and for derivation of the spectral Doppler signal prepared from the femoral artery. The investigation takes place with the ultrasonic system ATL UM-9 with the Tranducer type L 10-5.

The test animal receives an intravenous application of the test substance according to Example 8 or 19 and as a reference an application of a contrast agent produced according to WO 95/22994 (Example 12). The dose was 0.1 for all injections ml / kg body weight.  

The spectral Doppler signal is evaluated intensitometrically and against time applied. The resulting areas under the intensity-time curves are in the table 3 shown.

Table 3

Example 51

A beagle dog (female, 9.7 kg body weight) is anesthetized (inhalation anesthesia approx. 2/3 O₂; about 1/3 N₂O; 1.5-3% enflurane, spontaneous breathing) and Perfusion examination of the kidney prepared. The investigation will start with a Ultrasound system from HP (type Sonos 1000, 5 MHz) in color Doppler mode carried out. The experimental animal receives an intravenous application of a Agent according to the invention according to Example 22 (0.1 ml / kg body weight). After administration, the Perfusion representation of the organ compared to the representation before application clearly improved.

Example 52

A beagle dog (female, 9.7 kg body weight) is anesthetized (inhalation anesthesia approx. 2/3 O₂; about 1/3 N₂O; 1.5-3% enflurane, spontaneous breathing) and for sonographic Preparation of the abdominal aorta prepared. The examination is carried out with a Ultrasonic system of the brand ATL type UM9 with transducer C10-5 in harmonic B-  Fashion. The experimental animal receives an intravenous application of an invention Prepared according to Example 8 (dose 0.1 ml / kg body weight). Immediately after After the injection, the vascular volume is marked echogenically. Before the injection was the volume without enhancement.

Claims (16)

1. preparations for the preparation of a preparation for ultrasound diagnosis, consisting of a porous, solid, water-soluble matrix containing one low molecular weight scaffolders, a surfactant and a gas, the gas in the pores of the matrix. 2. Porous matrix according to claim 1 containing as a scaffold water-soluble scaffolders with a molecular weight (<15,000 Daltons) from the group of amino acids, polyamino acids, peptides, proteins, mono-, Di-, tri-, tetra-, oligo- and polymeric saccharides, as well as their derivatives, synthetic saccharides and their derivatives. 3. Porous matrix according to claim 1 or 2 containing L-glycine as a scaffold, L-alanine, L-valine, L-leucine, L-isoleucine, L-phenylalanine, L-proline, L- Hydroxyproline, L-serine, L-threonine, L-tryptophan, L-asparagine, L- Glutamine, L-arginine, L-histidine, glycyl-glycine, glycyl-glycyl-glycine, Glucose, galactose, fructose, mannose, sorbose, sucrose, lactose, Maltose, trehalose, gentiobiose, lactulose, turanose, maltotriose, melibiose, Melicosis, maltotetraose, maltopentaose, stachyose, arabinose, xylose, Ribose, dulcitol, xylitol, mannitol, ribitol, inositol, sorbitol, α, β, γ- Cyclodextrins, hydroxypropyl-β-cyclodextrin, dextran 8, dextrins and / or Ficoll. 4. Porous matrix according to claim 1 or 2 containing as a scaffold X-ray contrast media or contrast media for magnetic resonance imaging. 5. Porous matrix according to claim 4, containing iopromide as a scaffolding agent, Iotrolan, Iopamidol and / or Iohexol. 6. Porous matrix according to claim 4 containing Gd-DTPA as a scaffold (Gadopentetic acid), Gd-DTPA dimeglumine salt (Magnevist), Gd-EOB-DTPA (Gadoxetic acid, disodium salt) and / or Gadobutrol. 7. Porous matrix according to one of claims 1 to 6 containing as a surfactant water-soluble, nonionic surfactant, the surfactant portion of the matrix Is 0.01 to 10% (m / m).   8. Porous matrix according to one of claims 1 to 7 containing as a surfactant Sorbitan fatty acid esters, polyoxyethylene sorbitan fatty acid esters, Polyoxyethylene sorbitol fatty acid esters, polyoxyethylene fatty acid esters, Glycerol polyoxyethylene fatty acid esters, ethoxylated mono-, di-, triglycerides, which, if desired, can be partially hydrogenated and ethoxylated Mixtures of these, ethoxylated castor oils, ethoxylated phenols, Polyoxyethylene fatty alcohol ether, polyglycerol fatty acid ester, Sorbitan perfluoro fatty acid esters, polyoxyethylene sorbitan perfluoro fatty acid esters, Polyoxyethylene sorbitol perfluoro fatty acid esters, Polyoxyethylene perfluoro fatty acid esters, ethoxylated castor oils, Polyoxyethylene perfluorinated fatty alcohol ether, polyglycerol perfluorinated fatty acid ester, Polyoxyethylene polyoxypropylene polymers and / or fluoroalkyl Poly (ethylene oxide) alcohols. 9. Porous matrix according to one of claims 1 to 8 containing as a surfactant Surfactant with a perfluorinated hydrocarbon building block and / or with a Molecular weight <15,000 daltons. 10. Porous matrix according to claim 1 to 9 containing as gas nitrogen, oxygen, CO₂, air and fluorinated gaseous compounds. 11. Porous matrix according to claim 1 to 10 containing as gas tetrafluoroallene, Hexafluoro-1,3-butadiene, decafluorobutane, perfluoro-1-butenes, perfluoro-2-butenes, Perfluoro-2-butyne, octafluorocyclobutane, perfluorocyclobutene, Perfluorocyclopentane, perfluorodimethylamine, hexafluoroethane, Tetrafluorethylene, pentafluorothio (trifluor) methane, tetrafluoromethane, Perfluoropropane and / or perfluoropropylene. 12. Porous matrix according to claim 1 to 11 containing a gas as a perfluorinated Substance. 13. Ultrasound contrast medium generated from a porous matrix according to one of the Claims 1 to 12 containing water as the liquid carrier medium optionally with those customary in pharmaceutical technology Additives. 14. Ultrasound contrast medium according to claim 13 containing as a carrier medium physiological electrolyte solution, an aqueous solution of one or  polyhydric alcohols or an aqueous solution of a mono- or Disaccharides. 15. A kit for the production of an ultrasound contrast agent containing gas bubbles consisting of

• a) a first container provided with a closure which enables the removal of the contents under sterile conditions and is filled with the liquid suspension medium and
• b) a second container provided with a closure which allows the addition of the suspension medium under sterile conditions, filled with the porous matrix according to one of claims 1 to 12 and a gas, the volume of the second container being dimensioned such that the suspension medium of the first container completely in the second container.

16. A method for producing porous matrices according to one of claims 1 to 12, characterized in that first an aqueous solution of desired scaffold is produced, which if necessary gas bubble stabilizing surfactants are added, then the so prepared solution is freeze-dried after drying porous matrix is aerated with the desired gas.