DE1947896A1 - Pharmaceutical preparations and their manufacture - Google Patents

Description

DR. BERG DIPL.-ING. STAPF

8 MUNICH 2. Hl LBLESTRASSE 2O

Dr. Berg Dipl.-Ing. Stapf, 8 Munich 2, HilblestraBe 20 Your reference Our reference date

22 Sep 1969

Attorney File 18 852
Be / öch

L ¹ OREAL
Paris, France

"Pharmaceutical preparations and their manufacture" addendum to patent ...
Patent application P. 1667907.5

Inventor: Glaudine ßerrebi Georges Manoussos

The present invention relates to preparations that combat inflammation of cellular tissue, of aging and enable degeneration of skin tissues, anti-inflammatory or anti-inflammatory preparations, in particular

009838/2081

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against rheumatism, edema or burns ,, preparations to promote the healing process in wounds (scarring-promoting preparations), as well as procedures to their manufacture and their use as medicinal products.

The applicant has already proposed hair loss and beborrhoea with the help of cosmetic preparations to combat, which contain as active ingredients sterile extracts obtained by extracting the skin and tissues of the Umbilical cord from beef and sheep fetus with the help of a ion-containing solution can be obtained.

A salt, an acid or a base can be used as an ion-containing aqueous solution for the production of these active ingredient preparations or an ionizable organic compound such as a phenol dissolved in water can also be used. the Ion-containing aqueous solution can also be formed by a physiological serum solution.

Of the ionizable compounds that can be used to form the aqueous ion-containing solution, are mentioned:

- neutral courtship such as sodium chloride, potassium chloride, lithium chloride, Calcium chloride, magnesium chloride, in 0.1 M to 2.5 M and preferably 0.4 M solution,

- acidic mating like potassium or sodium phosphab in 0.1 W-

009838/208 1 _ ₃ _

BAO ORiQfNAL

Solution and a pH between 6 and 8 or sodium or potassium citrate in 0.1 M solution with a pH between 3 and 5i

- Acids, such as hydrochloric acid, sulfuric acid, acetic acid, in 0.5N Solution with a pH of 1.5 »

- Bases such as sodium hydroxide, potassium hydroxide, ammonia, oalcium * - or barium hydroxide, in 0.3 to 0.5 N solution with a pH from 8 to 9,

- organic compounds, such as urea, in 6 to 8M solution with a pH of 6 or phenol in a 2 solution.

According to one embodiment of the method, the skin of the fetus is first frozen, then grated and guided their enzymatic digestion in an ionic solution.

Such a digestion can advantageously with the help of enzymes from the group of proteolytic enzymes, such as proteinase, papain, pepsins and trypsin.

This digestion should preferably be effected without delipoiding. It essentially serves to cause the separation of protein substances in such a way that the mucopolysaccharides are released which, according to the invention, are to become the later extracts _{or the like}

009838/2081 -4-

ORIGINAL

The pulp obtained by the enzymatic digestion is then subjected to dialysis against distilled water, after it has previously been preferably subjected to centrifugation to separate out the solid parts.

After filtering the solution containing the active ingredients the latter is precipitated by adding absolute alcohol and sodium acetate.

The precipitate can then, for example by successive washing with alcohol, with a mixture of Alcohol and ether, and are purified with ether, and is then subjected to drying.

The active ingredient thus obtained is in the form of a white powder that is soluble in water and physiological serum solutions is.

According to a second embodiment of the method according to the invention, the skin and the umbilical cord are frozen, then treats them after they have been comminuted in an ionic solution, “like a physiological tissue, without previous delipoids. Then centrifugation is carried out and what is contained in the supernatant liquid is precipitated active ingredients with solvents such as absolute alcohol, acetone, etc. or with quaternary ammonium derivatives, such as cetyltrimethylammonium broraide or cetylpyridinium chloride the end.

0 0 9 8 3 8/2081 _ ₅ _

The precipitate is purified by washing with absolute alcohol, then with an alcohol-ether mixture, and obtained after drying, a white powder that is soluble in water and physiological solutions.

All phases of this process are carried out at a low temperature, for example a temperature of +4 ⁰ O.

The white powder, which is soluble in an aqueous medium and obtained by the above procedure, and the active ingredient which represents extracts of the skin cover is hereinafter referred to as "mucopolysaccharides according to the invention" .

The present invention has a cosmetic preparation suitable for local use, in particular against Cell tissue inflammation, copper rash (couperose), sun or Glacier fire and heat blisters and for activation wound healing or scar formation is the subject matter, the preparation in a suitable cosmetic carrier an effective content of mucopolysaccharides according to the Invention contains.

In a preferred embodiment, the cosmetic preparation according to the invention contains between 0.5 and 5 wt.%, And preferably between 1.5 and 3 wt.% Mucopolysaccharides.

009838/2081

-6-

In a preferred embodiment, the cosmetic preparations intended for use against cell tissue inflammation additionally contain enzymes which are known with regard to their effect on the metabolism of lipoids and mucopolysaccharides, such as, for example, enzymes from the group of hyaluronidases, lipases and thiomucases. These enzymes can be used in a concentration from 1 to 5 wt. % Be present.

The cosmetic preparations according to the invention can be presented in the form of solutions, gels or creams will.

»You can still use all conventional cosmetic aids, such as dyes, perfumes and / or penetrants (slide rail) contain.

The present invention likewise has a method of combating oellulitis, aging and degeneration of skin tissues, copper rash, sunburn and Heat flaking, the subject in which one applies the cosmetic preparations explained above to the infected Repeatedly applying skin surfaces and preferably massaging them at the same time.

The present invention also has a novel anti-inflammatory, anti-inflammatory, scarring-promoting, Skin tissue regenerating drug,

009838/2081

-9-

that also against varicosity, copper rash and burns acts, to the object containing the mucopolysaccharides according to the invention in a suitable carrier.

According to a first embodiment, this is explained above Medicinal product intended for administration by intramuscular injection and it is in this case as aqueous injectable solution containing the mucopolysaccharides are presented.

In a further form of presentation, the Medicines for external use administered and the mucopolysaccharides are in this case in a carrier, which is preferably formed by a cream or a gel, incorporated.

The medicament according to the invention can be administered in daily doses of 5 to 200 mg of mucopolysaccharides according to the invention per kg of weight of the patient being treated.

The therapeutic properties of the medicament according to the invention are essentially anti-inflammatory or anti-inflammatory, scar-forming and tissue-regenerating Properties,

The investigations carried out by the applicant show

009838/2081 ~ ⁸ ~

that the mucopolysaccharides according to the invention show no toxicity.

Furthermore, comparative studies allow the determination that the medicament according to the invention is anti-inflammatory or anti-inflammatory effectiveness and in particular has an anti-rheumatic activity similar to that of cortisone without the risk of harmful effects Secondary effects is significantly superior.

The examinations given below show the effectiveness and lack of toxicity of the drug according to the invention.

1. - Production of the mucopolysaccharide extracts from the Skin of the hind fetus

174-6 g of skin are removed under aseptic conditions of a bovine fetus.

The fetuses from which the removals are made are in gestation stages of 20 to 120 days, with the fetuses belong to one sex or the other without distinction.

Immediately after removal, the skin covers are frozen at a temperature of -20 ° C, then in the presence of a Saline solution (15 ml / g) in a U-Turrax TP 45/2 device

009838/2081 _9_

BADORiQINAL

minced at 10,000 rpm for about 15 minutes, these procedures being carried out under sterile conditions will.

The whole time it is chopped, it is cooled Whole with ice and carbonated snow to avoid any risk of spoilage.

In this case, the extraction salt solution is 0.1 M Phosphate buffer formed with a pH of 6.5. To the ground mass thus obtained are added:

300 mg papain
788 mg of cysteine chlorohydrate
18.6 mg ethylenediaminetetraacetic acid

to 100 ml of solution and digestion of the proteins is carried out 15 hours at 65 ° C.

During this phase, the sterility of the mileus is ensured by adding a small thymol crystal.

The resulting paste is treated with trichloroacetic acid with a final concentration of 7.5%. The pulp is then centrifuged for 30 minutes in a Jouan type 0 60 TS centrifuge at 4500 rpm and the supernatant is separated from ₀

This is then for 18 hours at 4 ⁰ C a dialysis

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subjected to distilled water, the dialysis membrane is made up of a cellulose tube.

The dialysate is then filtered through an adsorbent powder known as "Celite" is brought onto the market and the active "product is subsequently precipitated by adding 2 liters of absolute ethyl alcohol Un add 50 g sodium or potassium acetate per 1 solution.

The precipitation is successive through three the following washes with absolute alcohol, an alcohol-ether mixture and an ether ^ cleaned and then the drying subject.

All the process steps that have just been described are carried out at a maximum temperature of 4 ^° C. (except of course the enzymatic digestion)

The active product obtained is in the form of a white Powder which is soluble in water and in physiological serum solutions.

16 g of active ingredient in powder form are obtained, which is a yield of 9} 2 g per kg of skin.

The powder thus obtained, which forms the active ingredient, contains approximately 26 mg of hexosamines per gram of mucopolysaccharides.

According to another embodiment, one can use the same

00983872081

-11-

Carry out driving under the conditions described above, assuming 1881 g of umbilical cord as the starting material used by bovine fetus.

In this case, 4.25 g of active ingredient in powder form are obtained, which corresponds to 2.25 6 active substance per kg of umbilical cord.

In this! All the powder contains 185 mg of Hexosamine per g mucopolysaccharides.

2 - Production of mucopolysaccharide extracts from skin and umbilical cords from sheep embryos

The skins and umbilical cords are removed from sheep embryos regardless of gestation stage or gender. In this way, 500 g of skin cover and 1650 g of umbilical cords.

These products, removed under sterile conditions, are then frozen at a temperature of -20 C, then ground in a solution of 15 l of courtship solution per kg of tissue, without prior delipoiding, whereby the solution contains 9 g NaCl per liter of water.

The pulp is centrifuged for 90 minutes in a Wifug Xl centrifuge at 9000 rpm and the supernatant product is precipitated with 2 liters of ethyl alcohol and 50 g of sodium acetate per liter of solution. Is allowed the whole thing for 4 days at 4 ⁰ C rest

009838/2081 -12-

and collects the precipitate formed, which is cleaned by successive washing with absolute alcohol, then with a mixture of $ 0% alcohol and 50% ether, and then with ether alone.

The precipitate is then subjected to drying, whereby a white powder is obtained which is soluble in water and physiological serum.

All of these different process phases, which have just been described, are carried out at a temperature of at most 4 ^° C.

2.20 g of active ingredient are obtained if fetal skin is removed and 3.20 g of active ingredient in the case of using umbilical cords.

3 - Production of mucopolysaccharide extracts from skins

of child embryos

150 g of skin are removed from under aseptic conditions

4 to 5 month old bovine embryos.

This skin is pulverized after freezing in acetone, then stored at -20 ⁰ C dried. This pulp is then extracted with a 0.14 M sodium chloride solution in a ratio of 5 ml of solution per g of pulp, four on top of each other

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BATH ORIGINAL

carries out the following extractions under the same conditions, then stirred for 24 hours at a temperature of 4 0. The resulting solution is filtered through cheesecloth and then centrifuged for one hour at a speed of 6000 rpm. Then a 1.8 M perchloric acid solution is added in volume corresponding to half the volume of the saline extract in such a way that the free proteins are precipitated and are in the supernatant. After 4 minutes of standing at 4 ⁰ C was filtered through a Whatman filter Kr.4. The clear filtrate thus obtained is then added to half its volume of a 5 strength phosphotungstic acid solution in 2N hydrochloric acid. It is allowed to precipitate for one night, the temperature of 4 ^° C. being maintained. The precipitate is centrifuged and collected, washed successively three times with acetone and then dried under vacuum. The residue is absorbed by a 0.1 M sodium bicarbonate solution in a ratio of 3 rol / 5 g of fresh skin. Then 1 mg / 5 g of fresh skin dissolved trypsin in 1 ml of 0.001 N hydrochloric acid is added. After adding a small thymol crystal, the digestion is carried out at 37 ⁰ O for about 18 hours.

The proteins, which may be in combination with the mucopolysaccharides, are then precipitated by adding 3.1 H trichloroacetic acid at 1 ml / g of fresh skin. It is left to rest for one night at 4 ^° C. and then centrifuged for Λ hour at 4500 rpm. In the protruding one falls

009838/2081

-14-

Mucopolysaccharides by adding anhydrous sodium acetate (7> 5 millimoles per g of solution). Thereafter, the solution is twice its volume of absolute alcohol and can be a night at 4 ⁰ G flocculate, then centrifuged for 30 minutes at 4-500 rpm and successively washing the precipitation with alcohol, then with a mixture of alcohol and ether, and finally with Ether, after which one dries.

In this way, 135 mg of active ingredient in powder form are obtained, corresponding to a yield of approximately 0.5 g / kg fresh skin.

4 - manufacture; the mucopolysaccharide extracts from fiinderembr.yohäut en

Bovine embryo skins are removed under aseptic conditions, the skin in the presence of acetone, then from Ether finely grated and then the pulp is dried.

The product obtained is then suspended in 0.14 N sodium chloride solution, corresponding to 10 ml / g dry product. The mixture is stirred for 48 hours at 4 ⁰ G, then centrifuged for 90 minutes in the cold at a rate of 9'0OO rpm. The supernatant is then collected and dialyzed against running water for 24 hours. Distillation is carried out under reduced pressure at 50 ° C. in order to reduce the resulting solution to a quarter of the initial volume

0098387208t

-15-

focus. Lyophilization then takes place, after which the active ingredient according to the invention is obtained.

In another embodiment, the active ingredient contained in the supernatant layer can be precipitated in the same way by introducing a 1% Getyl pyridinium chloride solution and keeping the whole thing at 38 ⁰ O for 3 hours.

- Pharmacodynamic study of mucopolysaccharides the invention

I - effect on isolated organs

a) Action on the ileum of guinea pigs:

-Effect in the real sense-

The mucopolysaccharides show no effect on the smooth fiber at doses from 0.025 mg to 10 mg / 70 ml.

-Effect against acetylcholine-

The mucopolysaccharides neither potentiate nor inhibit the acetylcholine produced by 25 / Ug Ileum contraction.

The test doses vary in the range from 0.5 rog to 5 70 ml and the contact time between 30 seconds and 3 minutes th.

The action of the mucopolysaccharides was with and without

009838/2081 -16-

Washing of the organs determined.

-Effect against histamine-

Doses of mucopolysaccharides ranged from 0.25 mg to 40 mg have no effect whatsoever on the 1 / Ug histamine induced ileum contraction,

b) Effect on the duodenum of rats:

The mucopolysaccharides had no effect on the contraction caused by 25 / Ug acetylcholine Duodenum in rats. However, they can cause a slight decontraction after washing.

II - experiment on live rabbits

The effects of mucopolysaccharides on blood pressure, breathing and the electrocardiogram were examined and the chemical mediators like adrenaline, acetylcholine and histamine in rabbits.

The tests were carried out in white rabbits of both sexes with an average weight of 3 kg, with the help of a 10 # urethane solution in proportion of 10 ml / kg administered intravenously were anesthetized.

After the bleeding procedure, blood pressure is measured on the outer carotid with the help of a cannula

009838/2081 ~ ¹ ? -

is provided with a print head and a registration device.

Control of breathing is done in a similar way.

The electrocardiogram is triangular according to the EKG lead Dp right arm, left lower leg determined.

The cannulated external saphenous vein is used for intravenous injections.

The mucopolysaccharide extracts from the umbilical cords of calves in doses of 4, 8, 1Q, 2Q, 30, 40 and 50 mg / kg, intravenously administered do not lead to any disturbance of the normal pressure curve. In contrast, could under the effect repeated doses of mucopolysaccharides a delay in the respiratory amplitude can be observed.,

The mucopolysaccharide extracts from the embryo skin of calves, administered intravenously in low doses result in a moderate and transient hypotonic spike. At a At a dose of 30 mg / kg, sudden hypotension occurs, the intensity of which remains constant even with larger doses. but is short-lived.

The effect on breathing is different, it shows itself

00 9838 / 2.0 81

by slightly lowering the amplitude at low doses and by two-phase action at increased doses (50 mg / kg).

The mucopolysaccharide extracts from sheep embryo skin have no effect on arterial pressure up to the dose of 10 mg / kg. At 20 mg / kg one observes hypotension of weak intensity which is progressive but of fairly long duration.

An increase in the respiratory amplitude can be observed in the same way.

At doses of 10 to 20 mg / kg, the mucopolysaccharides have no effect on the blood pressure symptoms caused by adrenaline, acetylcholine and histamine.

At 25 nig / kg you can get a slight increase in adrenaline observe the hypotonic curve of adrenaline and a prolongation of the hypotension caused by acetylcholine. ·

In the electrocandogram, the doses of 10 and 20 mg / kg no change in both rhythm and amplitude.

Certain disturbances occur at 40 mg / kg. When the mucopolysaccharides are used locally in rabbits

009838/2081 -19-

no change in the blood pressure and respiratory processes is observed.

Finally, comparative studies between the mucopolysaccharides according to the invention and cortisone have shown that these have no effect on the percentage numerical values of the blood cells and on the blood count.

In this study, two groups of male rats weighing 250 g for eight days, one by intraperitoneal injection of the mucopolysaccharides according to the invention at a dose of 200 mg / kg, the other treated by intraperitoneal injection of cortisone at the same dose.

The percentage numerical values and the blood count were compared to a group of normal control rats on the ninth day compared

Control animals mucopolysaccharide cortisone

Blood cells 16 500 16 520 8 600

Furthermore, the decrease in the number of eosinophils and lymphocytes, as well as the increase in polynuclear cells

-20- 009 838/2081

after treatment with mucopolysaccharides ^ compared to the with cortisone means less pronounced.

The mucopolysaccharides according to the invention accordingly have little or no secondary effects that are responsible for their Use would be harmful.

II - Study of the effectiveness against oellulitis of the Mucopolysaccharides of the invention

These studies were carried out on "Wistar" rats, a number of which have undergone ovarian extirpation. Indeed appears after ovarian extirpation in rats a lipoid and water storage which is very similar to the manifestation that in women from cell tissue inflammation (oellulitis: in French Literature common name for various clinical pictures of the face, lower extremities or as General condition) occurs. This effect is generally seen after a period of 2 to 3 months in rats obtained after ovarian extirpation.

The experiment required three groups of 10 animals each same parentage,

a) the first group are normal female control animals that are not ovarian extirpated,

b) the second group are ovarian extirpated females

009838/2081 -21-

Control animals,

c) in the third group, the active substance is used, the rats of this group having ovarian extirpation in the same manner.

All rats of groups b) and c) were ovarian-extirpated on the same day and in the experiment reported here an effect analogous to that of cellulitis in all rats two and a half months after ovarian extirpation observed.

The experiment was carried out as follows:

- the first group was not subjected to any treatment,

- the second group was given 0.5 ml daily for 12 days treated with distilled water per animal

- and the third group was treated in the same way daily for 12 days with 0.5 ml of active ingredient per animal, with the active ingredient is an extract solution from embryonic organs

in distilled water was made according to that described above Production method No. 1 was obtained and its concentration was 2.5 g of dry extract / 100 ml of solution.

The second and third groups are administered on a back surface of about 3 cm that before was sheared.

—22—

009838/2081

The day after the last treatment, all batteries were weighed and killed.

The skin removals appropriate to the areas of application are determined with regard to water and lipoid weight, whereby the following mean results were obtained

State of the animals modification
the Ge
weight Water
stop Lipoid
stop not ovarian
extirpated + 9g 290 136 Ovaries extirpated
(Control) + 9 g 329 156 Ovaries extinct
treated + 5g 285 110

This experiment was carried out under the same conditions with a repeated in another group of animals. The results obtained have been confirmed.

In view of the experimental results, it appears that the application of embryonic organ extracts to rat skin a reduction in lipoid storage of the skin parts and also causes a reduction in their water retention.

You can also tell that it is not a 0098 387 2081 -23-

Dehydration of the skin is a regulatory effect the water content, because the water content of the normal untreated and treated ovaries stipulated Rats can be seen lying together.

III - Investigation of. Ultraviolet erythema

radiation

20 rabbits "Alinos Bouscat" weighing 2.5 to 3 kg were first shaved to 1/10 mm in the area of the sternum and abdominal region, then with ultraviolet rays for 15 minutes irradiated, the irradiation being carried out with a lamp of 500 watts at a distance of 35 cm from the animal skin and the irradiation

ρ
was made even on areas of 3 cm, each area being determined by a mask. After 48 hours have elapsed after the irradiation, 2 ml of active ingredient solution is applied to the areas damaged by these irradiations in 10 rabbits, this consisting of embryonic tissue extracts obtained according to the above-described preparation form no. 1, with a concentration of 2% - Dry extract.

4 to 7 days are necessary for scarring in the animals treated with these preparations, while the control animals Need 10 to 15 days for scarring.

From the experiment it can be seen that the 009838/2081

- 24 active ingredients cut healing time by approximately $ 50.

Similar results are obtained with preparations in the form of ointments, these containing the following proportions:

Paraffin oil 30 g

Stearic acid "3 g

Triethanolamine · 0.2 g

Tween 60 (sorbitan-oxethylated mono-

stearate) '2g

Arlacel 165 (self-emulsifiable

Glycerine monostearate) 6 g

Bactericid 0.3 g

Extracts from embryonic tissues, which are produced according to the manufacturing process described above method no. 1 obtained 1 g

Water to 100 g

IV - Study of the anti-inflammatory or anti-inflammatory effectiveness of the preparations of the invention

1 - The anti-inflammatory activity of the mucopolysaccharide-containing preparations of the invention examined with the help of an experimental kaolin edema.

For this purpose, female rats of 150 g from the same breed were divided into groups of 10 animals each.

The edema is obtained by injecting 0.1 ml of a sterile

009838/2081 " ²⁵ ~

bus boarding from 10 $ kaolin under the t> ohle of the left Paw of the animal provoked.

In each experiment, the first group of animals serves as a control group. The difference between the weights on the left rear paw of each animal in which the edema was provoked and the rear right paw that is intact serves as a measure of the intensity of the edema.

The other animal groups undergo preventive or curative treatments on the one hand using mucopolysaccharides subject to the invention and, on the other hand, by means of cortisone.

These different treatments resulted in a reduction in what was provoked in the animal's left rear paw Result in edema.

The reduction in % in the weight difference between the left rear paw and right rear paw of the animals of each group relative to the weight difference between the left rear paw and right rear paw of the control animals enables the effectiveness of the various treatments tested to be measured.

The anti-inflammatory effectiveness is 3 hours after the local kaolin injection measured and the animal through

009838/2 081 _ ₂₆ _

Bleeding killed. You then remove the two back paws and the difference in weight between the two Paws is a measure of the intensity of the edema.

The treatment with mucopolysaccharides of the invention can at a different time with regard to the kaolin injection take place, whereby one can observe the curative or preventive effect.

The mucopolysaccharide solutions of the invention are injected in the ratio of 1 ml / 100 g body weight.

In the case of skin traces, these are done with help a solution of the mucopolysaccharides, the concentration of which is 50 mg per ml. Every application receives the Animal 0.5 ml of this solution.

In the majority of cases, the anti-inflammatory or anti-inflammatory efficacy of the mucopolysaccharides was of the invention equivalent to that of cortisone.

A. Mucopolysaccharide extracts from calves

The mucopolysaccharides of the invention were in one case Extracts from embryo skin and in the other from umbilical cords.

a) preventive treatment:

The mucopolysaccharides are administered in three doses of 200 mg / kg in 24 hours before the local kaolin injection,

009838/2081 _ _2? _

taking the first dose 24 hours prior to treatment with Kaolin is given. The second and third doses are given 1 hour before the kaolin, respectively.

The results obtained are shown in the table below:

200 mg / kg embryonic skin

intraperi- - umbilical cord

tonal '- Cortisone $ 59

200 mg / kg - embryonic skin
subcutaneous cortisone

- 0.5 ml - embryo skin

Hautbe- - umbilical cord 20 #

plot

b) Medical treatment:

The animals received three doses in 200 mg / kg extract of Mucopolysaochariden according to the invention, one hour, 18 Hours and 23 hours after kaolin application.

The animals are sacrificed 1 hour after the third dose. The following results were obtained:

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200 mg / kg - embryo skin
subcutaneous - cortisone

0.5 ml skin - embryo skin
treatment - cortisone

B - Mutton mucopolysaccharide extracts

a) Short-term preventive treatment by intraperitoneal Injection of various doses.

As in the previous studies, the effectiveness of the mucopolysaccharides of the invention with the. from Compared cortisone and summarized the results observed below:

dose Embryo skin Cortisone 5 mg Λ CJOL
I OyO . 8th% 25 mg 21% 50 mg 29% 100 mg 200 mg 8 6 *. 78%, 87 * 4-5 $, 51 #, 28 # ⁺ intraperitoneal
injection

b) Short-term preventive treatment by local application 009838/2081 -29-

To the effectiveness of the mucopolysaccharides according to the invention To compare, a hydrocortx sonic ointment was used, while the mucopolysaccharides were buffered in a Solution have been dissolved.

The following results were obtained:

- 0.5 ml - erabryo skin

- 500 mg pomade

with 2.5 # cortisone

The anti-inflammatory effectiveness was also continued with the experimental kaolin edema, but in had the adrenal glands had been removed.

The effect of the mucopolysaccharides was found under test conditions which corresponded to the previous ones of the invention in rats that had their adrenal glands removed were unchanged again. The mucopolysaccharides do not act via the adrenal glands in the sense of an administration of cortisone. Their effect is completely independent of this.

2- The anti-inflammatory or anti-inflammatory efficacy of the mucopolysaccharides of the invention has continued

009838/2 081 ~ ³ ° "

studied in other experimental edema.

a) Experimental edema with bradykinin

This edema was injected under the sole of the foot 0.1 ml of a solution with 0.1 mg / ml synthetic bradykinin ("calbiochem") provoked. In this attempt a preventive treatment carried out 1 hour in advance and the Animals were sacrificed 15 minutes after the appearance of the edema,

- 200 mg / kg embryo skin

intraperitoneally - cortisone

b) Experimental edema with carraghenin

Experimental edema with carraghenin manifests itself most similarly to rheumatic diseases. This edema provoked under conditions similar to those obtained with the help of kaolin, an injection of 0.1 ml of 2 # carraghenin solution under the sole of the animal.

- 200 mg / kg - embryo skin $ 68

intraperitoneally - cortisone

V - Investigation of the capillary permeability 'of the preparations of the invention

For this purpose the rats are given 009838/2081 on the day before the experiment

-31-

shorn at her back. An intravenous injection of 1 ml of 1 # igera "Prontamine Sky Blue" is then administered. Five minutes after this injection, the rats received three intradermal injections of 0.1 ml on the left side 1 mg / ml histamine solution. You then set the time up to to the appearance of the dye at the level of the intradermal papula. The same rats are then given intraperitoneal means Injection of the mucopolysaccharides of the invention at a dose of 200 mg / kg. After 30 minutes the right side of the rats the same treatment as the left side. One then poses as in the previous case the times of color appearance fixed.

Time left side right side

(Control) (treated) color appearance

5 i

The mucopolysaccharides accordingly have a distinct Effect on capillary permeability.

VI - Investigation of the scar-forming effectiveness in wounds, the healing of which is experimentally disturbed by cortisone.

Epithelial tissue wounds are applied to rabbits' ears using punch pliers. The scarring is caused by daily intraperitoneal injection of a cortisone solution

009838/20 81 ~ ³² ~

25 mg / ml in the ratio of 25 mg / kg body weight during 4 days delayed.

Daily use of a solution of mucopolysaccharides for 15 days of the invention with a concentration of 50 mg / ml in the ratio of 0.1 ml per wound, brings the Scarring curve, especially after the tenth day, to normal Sequence.

In the case of an injection of the mucopolysaccharides by means of intraperitoneal Administration of 25 mg / kg will return the scarring curve to normal after the fourth day.

Examples of the preparations of the invention.

example 1

A solution is prepared for injection at the site of inflammation, using the at the moment of use Mix the contents of the following two ampoules:

1. Ampoule: 5 mg or 10 mg of mucopolysaccharides, in turn were obtained by the manufacturing method 1.

2. Ampoule: 5 ml sterile, non-feverish pH 7 buffered solvent *

The composition of this solvent is, per 1000 ml Water, 1 g monopotassium phosphate, 1 g sodium bicarbonate and

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9 g sodium chloride «

25th S. 8th S. 10 S. ο, 2 pp 2 S. 100 S.

Example 2

A scar-forming aerosol foam is produced, wherein the following components in the aerosol container

mixes:

Cross-linked polyacrylic acid, marketed under the name
"OARBOPQL- 934"

Magnesia oxylauryl sulfate glycerin ammonia

Mucopolysaccharides produced by process no.1

Water on

88 g of the solution described above are taken, placing it in an aerosol container with 12 g of difluorodichloromethane conditioned.

Example 3

A powerful cream is produced against copper rash (Kupferose) and varicose veins, whereby the following ingredients are mixed, works:

Mucopolysaccharides obtained by Production Method 1, 2 g

oxethylated cetostearyl alcohol 7 S

009838/2081

Silicone oil 1 g

Diethylene glycol stearate 6g

Methyl parahydroxybenzoate preservatives, put on the market under the name "NIPAGINE" 0.10 g

Propyl parahydroxybenzoate preservatives, put on the market under the name "JpPASOL" 0.10 g

Water to 100 ecm

Example 4

A penetrating gel against cellulite is made from the following ingredients:

lyophilized extract according to the manufacturing process 1 3 g

Perfume in sufficient quantity

Gel Garbopol pH 8 to 1000 g

The Garbopol gel is prepared in such a way that Carbopol 94-0 is distilled in a ratio of 1 g to 100 ml Dissolve water at 70 ° C until a homogeneous gel is obtained. After cooling, the gel is brought to pH 8 with an ammonia solution by triturating it with a mortar homogenize it well and then add sodium merthiolate parts / 10,000.

-35 -00 983 8/2081

Claims (11)

New patent claims 1. Cosmetic or pharmaceutical preparation which is used for the local treatment, in particular of cellulite, Pits, copper rash, sunburn, and heat flaking is suitable or anti-rheumatic, anti-inflammatory or anti-inflammatory, wound-healing, i.e. scarring-promoting, skin tissue-regenerating medicinal preparation, which also against varicose veins, copper rash and burns is effective, characterized in that it is sterile in a suitable cosmetic carrier _{x / x} 009838/2081 Contains extracts obtained by extraction by means of an ion-containing solution from the skin and / or the umbilical cord tissues obtained from beef or sheep fetuses. 2. Cosmetic preparation according to claim .1, characterized in that it contains between 0.5 and 5% active ingredient. 3. Cosmetic preparation effective against cellulite according to claim 1, characterized in that they also have enzymes with an effect on the metabolism of Contains lipids and mucopolysaccharides. 4. Cosmetic preparation according to claim 3, characterized in that the enzymes hyaluronidases, lipases and / or are thiomucases. 5. Cosmetic preparation according to one of claims 3 and 4, characterized in that the en-zyme in a concentration of 1 to 5 wt -.% Are present. 6. Cosmetic preparation according to one of claims 1 to 5j, characterized in that it is in the form of a Lotion, gel or cream is present. 00 983 8/208 1 - "* 7. Cosmetic preparation according to claim 6, characterized in that it cosmetic adjuvants, such as Dyes, perfumes or penetration-inducing agents contains. 8. A pharmaceutical preparation according to claim 1, characterized in that it is in the form of a through the skin the preparation to be administered is present. 9. A pharmaceutical preparation according to claim 1, characterized in that it is in the form of an intramuscular Injection is the preparation to be administered. 10. A pharmaceutical preparation according to claim 1, characterized in that it is in the form of a cream or one Gels present. 11. Pharmaceutical composition according to one of claims 1 and 8 to 10, characterized in that it has a concentration of 0.5 to 5 wt -% contains mucopolysaccharides.. 009838/2081