Ultrasonic contrast agents, process for producing them and their use as diagnostic and therapeutic agents.

The present particle according to preamble a process for its preparation and invention relates to the micro of claim 1, to its use as the^![people for diagnosis and therapy. !

h Contrasts are known for echoes | Cardiac disorders can be achieved by peripheral injection of solutions containing fine gas bubbles (Roelandt J, ultrasound ed. 8: 471-492). These gas bubbles are dissolved in physiologically compatible solutions, for example by shaking or shaking or by adding carbon dioxide. :

I r \) «Λ

<img file="PT89635B_D0001.tif" />

the davia are not normalized in cs number lathes

Ç;

the bone and its period are raised to the erythrocytes, whereby the carillary or pulmonary organs such as the left ventricle, liver, kidneys or spleen. Furthermore, they are not suitable for quantification as an ultrasonic echo which they provide consists of ez or the color:

of the other various processes, such as mode formation is not in these blisters, coalescence and can be obtained for example by transit times with the aid of ultrasonic contrast by measuring the contrast in the myocardium. This requires agents whose dispersed bodies are not subject to their own kinetics.

course against ihirt, .. 'to 1.1ad, Ultrasonic Lackscatter from Contrast-producin: i Collagen ..iicro spheres, Ultrasonic Imaging 2: 66-67, 1930).

on the other hand higher density solutions are used; as ultrasonic contrast agents (C-phir, fcV / hirt,

I hlad, Aqueous Solutions as a Potential Ultrasonic Contrast Agents, Ultrasonic Imaging 1: 265-279, 1979, is just like that Tgler Ophir, Aaklad, In-vivo Snhancemente of Ultrasonic Image iu.iiinancs Soon, Ultrasonic Imagination 5: 525-329, 1981). Also, the use of emulsions as ultrasonic contrast agents is known (J. et al., André et al., Ultrasonic Enhancement of Llyocardial Infarction, Fluorocarbon Compounds in Dogs, Am. J. Carciol 54;

206-210.1984). I

Globally, gas-free contrast agents have only poor efficiency. Gas-containing preparations are unlikely to have only slight stability in vivo. In addition 2 hunted in venous sis

<img file="PT89635B_D0002.tif" />

which contain small bubbles d bubble

<img file="PT89635B_D0003.tif" />

injures asq · 'against

235 and

Patents no. A2 125 s teraa of arterial vessels capillary tubes of the lungs producing the effect

934 describes hollow ultrasound contrast voids, gas hollows. However with none of the ultrasound contrast agents known to date, it has sufficient signal intensity through selective concentration following an iv dose. Therefore, it is not possible at the present time to achieve considerably higher determinable and orally measurable quantification and can be concentrated intracellularly thereafter in the liver or spleen.

This is achieved only with microparticles consisting of a biodegradable polymer synthesized by a fluid with a boiling point of acidic acids. -, ô -, f Polyester or is -hydroxycarbonic, polyalkyl cyanoacrylates, rolia minoacids, polyamides, saccharide polyacrylates or rolo-orthoxysers are referred to as biodegradable polymers.

<img file="PT89635B_D0004.tif" />

torics.

The following compounds have been shown to be particularly suitable:

Polylactic acid,

Poly-caprolactone, lactic acid docolymers and β-lactic acid or β-acetone,

Polyhydroxybutyric acid, polyhydroxy valeric acid, hydroxybutyric and hydroxy valeric acid polymers,

Glutamic acid and / or lysine polymers,

Polydioxanone: Polymers or copolymers of amino acids and / or terephthalic acid, italic acid or setacic acid; Polyacryl dextran,

I polyacrylic starch,

Polyacrylamine,

Polyurethane, | ; ?O.' i-t = r,, Polyethoxy, 1 Polyaminocriazole or; polyalkyl cyanoacrylate

The microparticles may also contain starch or starch derivatives. Amyloses have been shown to be particularly suitable since such acid derivatives have excellent water solubility and capacity. to provide inclusion compounds.

Particularly suitable amylases are cyclodextrins and their derivatives, for example, 9, 9 and cyclodextrin.

Microparticles contain gases and / or fluids with a boiling point of less than COO.^JC in one form; free or on. Likewise it is also possible to use! uses gas / fluid mixture in the last-only contrast agents

<img file="PT89635B_D0005.tif" />

<img file="PT89635B_D0006.tif" />

The gases used may be, for example, air, nitrogen, inert gases, hydrogen, carbon dioxide, ammonia, oxygen, methane, ethane, propane, butane, ethylene or other hydrocarbons or mixtures thereof.

Preferred or incorporable fluids are:

1,1-diol ethylene,

2-methyl-2-butene, i

J isopropyl chloride,

2-methyl-1,3-butadiene,

P 2-butyne,

2-methyl-1-butene, dibromo-difluoromethane, iurane,

| 3-methyl-1-butene,

Isopentane, diethyl ether, 3,3-dimotyl-1-butine,

H . .

Cim.-ethyl-acetone, single oropylene,

N, N-ethyl methyl amine, bromine, methane, n-ethyl dimethyl amine, methylene chloride, pentane, cyclopentane,

I 2,3-pentadiene, cyclo-error or mixtures thereof.

Microparticles may also contain substances with low vapor pressure and / or low boiling points, in particular ethereal oils.

It is particularly advantageous to coat

<img file="PT89635B_D0007.tif" />

1'-I try. consequently the microparticles may. be surrounded by oils, fats and / or surface-active substances and placed in suspension in an aqueous medium.

It is particularly advantageous to involve!

| ' microparticles consisting of amylose by a matrix,

I more particularly by a polymeric structure .:

I * “i

Physiological isotonia may be adjusted. ¹ by addition of osmotically active substances such as

¹ the cooking salt, galactose, glucose, iructose. '

An advantageous process for preparing _: ultrasonic contrast agents according to I; The present invention consists of dissolving a polymer or co | polymer in one or more organic solvents which are not

I ₍ ; miscible with water, followed by emulsification and water, possibly with the addition of another solvent and then filtering. drying the obtained emulsion.

i

Another process is to dissolve a polymer or copolymer in one or more solvents which do not contain gas bubbles after precipitation or emulsification in water if necessary with the addition of another solvent or another. polymer and then filtered

<td>| if the suspension or emulsion</td><td>and</td><td>if necessary</td><td>dries up.</td>

<td>C freeze drying process</td><td>also</td><td>and appropriate as</td><td>c rc cesso</td>

<td>termination. 1</td><td></td><td></td><td></td>

<td>The</td><td>product</td><td>s obtained can</td><td>be with</td>

advantage, finely ground,.

In such processes the solvents may be, for example, furan, pentane, acetone, dioxene, ethyl acetate, xylol, methylene chloride, cyclohexane or hexane or mixtures thereof. It is also possible to add emulsifiers to the emulsion.

!

'Smoke another variant of the process. preparation instead of polymers monomers are used as

<img file="PT89635B_D0008.tif" />

starting product from which the polymer is formed. In accordance with this process, a naked monomer or various organic solvents are dissolved and then emulsified in 5-50 parts of water or 0.01-0.1 N hydrochloric acid, if necessary with the addition of emulsifiers or - Buffer substances at a temperature below the boiling point of the organic solvent, after which an emulsion of 0.2 / -20% of a second monomer is added to this emulsion or, if necessary, a solution of a substance which increases the pH value, and then dry if necessary.

In another method of preparation, a monomer is dissolved or dispersed in one or more fluids containing gas bubbles, if necessary with the addition of emulsifiers or buffer substances. If necessary add to this solution or dispersion a solution between 0.2 / - 20/2 of a second monomer or a substance, or increase the pH value in a dissolved or gaseous form, and then dry if necessary.

By way of example, terephthaloyl or sebacoyl chloride or cyanoacrylic acid ester as the first monomer are used as the first monomer, and L-lysine as the second monomer and as an organic solvent, for example 2-methyl-1,3-butadiene is used. dioxane, methylene chloride, toluene or cyclohexane.

According to an additional process, ultrasonic contrast agents are prepared by producing gas bubbles in a 0.5-10% aqueous solution or dispersion of a monomer containing, if necessary, additives such as emulsifiers (0.01 - 5%) or emulsifying ouase (0.1 - 5u) and then adding a substance which provides cross-linked bonds and / or a reaction initiator.

The ultrasonic contrast agents described above can be used in both diagnostic and therapeutic processes.

<img file="PT89635B_D0009.tif" />

The application of the agents is done by injection, for example.

The present invention will be further illustrated by the following examples:

: EXAMPLE 1:

ι ------------------------------------------------- -------------------------------------------------- -------------------------------------------------- ----! 500 mg of polylactide was dissolved in 4 ml of furan and 0.6 ml of cyclohexane, and this solution was emulsified in 40 ml of a 0.1% solution of polyoxyethylene polyoxypropylene polymer of molecular weight. ..

^! 12,000 (Pluronic © 127), maintaining the temperature below 15 ° C during emulsification. Then raise the temperature slowly to evaporate the organic solvent. The resulting suspension is then lyophilized.

EXAMPLE 2:

500 mg of cyano-acrylic acid ethyl ester was dissolved in 1 ml of furan and this solution was dissolved in 10 ml of 0.1 N hydrochloric acid containing 1% polyoxyethylene polyoxypropylene polymer with a molecular weight of 12,000 (Pluronic (127), maintaining the temperature below 15 ° C during emulsification. At the end of the polymerization the resulting suspension is lyophilized.

EXAMPLE 3:

200 ml of cyanoacrylic acid butyl ester was dissolved in 0.4 ml of isoprene and emulsified to 30 ml of 0.01 N hydrochloric acid containing 1% (by weight molecular weight polyoxyethylene polyoxypropylene roller). of 3,350 (Pluronic ™ 68), keeping the temperature below 10 ° C during emulsification.At the end of the polymerization the suspension with 0.1 N NaOH was neutralized and the isotony with sodium chloride adjusted.

<img file="PT89635B_D0010.tif" />

EXAMPLE 4: ί

400 mg of the cyano-acrylic acid butyl ester and 0.14 ml of methylene chloride were dissolved in 60 ml of 0.01 M hydrochloric acid containing 1% polyoxyethylene polyoxypropylene polymer with a molecular weight of 12,000 (Pluronic 0 F 127); keeping the temperature below 10 ° C during emulsification. At the end of the polymerization the suspension was neutralized^!with 0.1 N sodium hydroxide solution and isotonia was adjusted with sodium chloride.

EXAMPLE 5:

400 mg of polycaprolactone was dissolved in one ml of furan and 0.3 ml of cyclohexane and emulsion; 60% of 1% polyoxyethylene polyoxyoxypylene polymer having a molecular weight of 12000 (Pluronium 127) was maintained at a temperature below 15%. The temperature was then slowly raised to evaporate the organic solvent. The resulting suspension was then lyophilized. !

EXAMPLE 6;

400 mg of terephthalic acid dichloride was dissolved in 2 ml of furan and then 50 ml of a 3: 7 sodium carbonate solution containing 0.1: of a polyoxyethylene polyoxypropylene polymer of a weight molecular weight 12,000 (Pluronic 0.127). After addition of 60 mg of L-lysine dissolved in 5 ml of 0.1 µl Pluronic E 127, the microcapsules were centrifuged and washed several times with a 0.1% solution of Pluronic F 127. Prior to use, the suspension isotonia was adjusted with sodium chloride.

EXAMPLE 7:

Β-cyclodextrin / isopentane inclusion compound;

I

<img file="PT89635B_D0011.tif" />

'Ο ι

It was cooled to 10 0 10C ml of one! saturated solution of α-f) -cyclodextrin (1,3) -) and mixed with 3 ml of isopentane. The resulting poorly soluble complex was precipitated with constant agitation in an ultrasonic bath. The deposit was obtained in crystalline form α! through freeze drying and filtration. Isopentane content according to GC calculations (Gas chromatography): 0.25b.

EXEilP10 8; i

Inclusion compoundz5-cyclodextrin / 2-methyl-2-butene;

100 ml of a saturated solution of β-cyclodextrin (1.8%) was cooled to 10 ° C and mixed with 5 ml of 2-methyl-2-butene. The resulting poorly soluble complex was precipitated with constant agitation in an ultrasonic bath. The deposit in crystalline form was obtained by lyophilization and filtration.

i

I EXEhPLO 9:

Cyclodextrin / 2-methyl-1-butene inclusion compound:

100 ml of a saturated solution of cyclocextrin (1.8) (0) was cooled to 10 ° C and mixed with 5 ml of 2-methyl-1-butene. The resulting poorly soluble complex was precipitated with constant agitation in an ultrasonic bath. The deposit was obtained in crystalline form! by lyophilization and filtration. 2-Methyl-1-butene content according to GC calculations; 0.82 /.

I

EXE1IPL0 10:

Α-9-Cyclodextrin / Isoprene Inclusion Compound:

100 ml of a saturated β-cyclodextrin solution (1.8%) was cooled to 10 ° C and mixed with 5 ml of isoprene. The resulting complex is hardly

<img file="PT89635B_D0012.tif" />

soluble was precipitated with constant stirring well; ultrasonic. Deposition in crystalline form was obtained by lyophilization and filtration. Isoprene content according to

I GC calculations: 1.0.

EXAMPLE 11;

4) -cyclodextrin / isopropyl chloride inclusion compound

100 ml of a saturated β-cyclodextrin solution (1.8%) were cooled to 1 ° C and mixed with 3 ml of isopropyl chloride. The resulting poorly soluble complex was precipitated with constant agitation in an ultrasonic bath. The deposit in crystalline form was obtained by lyophilization and filtration. Isopropyl chloride content according to GC calculations: 0.5%.

EXAMPLE 12:

4-Cyclodextrine / opentane inclusion compound:

i | 100 ml saturated β-cyclodextrin solution (1.8%) was cooled to 10 ° C and mixed with isopentane 3 ml. The resulting poorly soluble complex was precipitated with constant agitation in an ultrasonic bath. The deposit in crystalline form was obtained by lyophilization and filtration.

i

EXAMPLE 13;

Xenon / cA-cyclohexocyte inclusion compound:

I

I; 100 ml of a saturated decA-cyclodextrin solution (about 12%) was incubated under. a pressure of 7 xenon atmospheres for 7 days at room temperature in a 200 cc autoclave. The crystalline aletum was aspirated, washed with cold water and dried with calcium chloride in the desiccator.

<img file="PT89635B_D0013.tif" />

14:

Carbon Dioxide Inclusion Compound / * - cyclodextrin:

100 ml of a saturated S-cyclodextrin solution (about 12%) was incubated for 7 days at room temperature under a pressure of 7 atmospheres of carbon dioxide in an autoclave of 20 ° C. cold water and

The crystalline adduct was removed, washed with dried with calcium chloride in the desiccator.

EXAMPLE 15:

Hydroxypropyl-z3-cyclodextrin inclusion compound:

15% 20% hydroxypropyl-7Î ± -cyclodextrin was mixed with 10Â ° C, a solution of the resulting complex of ultrasound was sonicated and then incubated.

remained in for for isopentane minutes hours.

EXAMPLE 16:

Droxypropyl-3-cyclo dextrin compound;

isoprene inclusion / hiMixed ml

20% hydroxypropyl-β-cyclodextrin at 10 ° C was sonicated in an ultrasonic bath and then the resulting complex was incubated and partially precipitated into a solution of ml of rsonreno during during solution and a white deposit.

EXAMPLE 17:

Furan / hydroxypropyl-e?) -Cyclodextrin inclusion compound:

15 ml of a stock solution was mixed.

20% hydroxypropyl cyclodextrin with 2 ml of furan at

<img file="PT89635B_D0014.tif" />

'! At 10 ° C, it was sonicated for 3 minutes in an ultrasonic bath and then incubated for 26 hours. *

The resulting complex remained partially in solution and partially precipitated as a white deposit.

Hey j ''

H EXAMPLE 18: '| Isopentane Inclusion Compound / J / J

I-dextrin cycle:

20 ml of a saturated cyclodextrin solution was mixed with 1 ml of isopentane and sonicated for 3 minutes in an ultrasonic bath. The resulting complex hardly obtained. ; ) was filtered off and dried with calcium chloride.

i ι

EXAMPLE 19:

Isoprene Inclusion Compound / (A-cyclodex trine:

20 ml of a saturated solution was mixed. diclodextrin with 1 ml isoprenc and sonicated for 3 minutes in an ultrasonic bath. The resulting complex, which was difficult to soluble by filtration, was obtained and dried with calcium chloride.

! EXAMPLE 20: I

-------------------------------------------------- -------------------------------------------------- -------------------------------------------------- ---- | i

Furan Inclusion Compound / cA-ci-1 | clodextrin;

í

20 ml of a saturated Î ± -cyclodextrin solution with 1 ml of furan was mixed and sonicated for 5 minutes in an ultrasonic bath. single. The resulting poorly soluble complex was obtained by filtration and dried with calcium chloride.

EXAMPLE 21;

Drop by drop added 4 g of eu13 li

I

<img file="PT89635B_D0015.tif" />

i

<img file="PT89635B_D0016.tif" />

oaliptol to 100 ml of a saturated solution (5 ° C) in an incubation chamber while sonicating and then sonication continued for an additional 30 minutes. The incubation chamber was then shaken in a cool closed container

<td>during</td><td>48 hours</td>

<td>I'm going</td><td>with ethanol</td>

<td>filied</td><td>- S 6.</td>

<td>EXE.IPLO</td><td> 22:</td>

The precipitate, the liquid precipitated, the liquid precipitated, and the liquid precipitated.

100 ml of a 2 g saturated de-cyclodextrin solution - iraniol ab C for 4 hours and then incubated for 24 hours at 5 ° C. The resulting precipitate was filtered off, washed with cold ethanol, frozen in nitrogen and lyophilized.

following statement is airable to Examples 7-22:

In physiological cooking, crystalline deposit was absorbed by appropriate acidity, preferably glucose salt or nino solution thus ready for injection.

staying