DE1467908A1 - Process for obtaining pacenta material - Google Patents

Description

PATENTANWi DR. LOTTERHOS - DR.-ING. LOTTERHOS

«000 PRANHrUMT fMAIN) 1 Λ K 7 Q D P

AKNASTRASSE 19 Itü ί OUO

FEk.! SPEAKER; (0611) 55S061
TELEGRAMS: LOMOSAPATENT
LANDESZENTRALBANK 4,951
DRESDNER BANK FFM., No. 524749
POSTAL CHECK. ACCOUNT FfM. 1661

^γ / Χ FRANKFURT (MAIN), 17, llär * 1964

Louis W. Granirer, 9o-36 149th Street »
Jamaica 35, New York.

Process for the extraction of piacenta material

The invention relates to a method for the production of pharmaceuticals valuable material from the placenta that can be used in both human and veterinary medicine

According to the method of US Pat. No. 2,907,695, it is recovered of pharmaceutically valuable material from the human Placenta the finely comminuted placenta is extracted with acidic, aqueous ethyl or methyl alcohol and extracted from the obtained Extract with a base the pharmaceutically valuable material rlacentamaterial man - like "

The structural formula of this ίlacenta material is not known However, it is a crystalline magnesium phosphate complex, the analytical data of which is contained in U.S. Patent 2,907,695 are. Further analysis data are given below

it was found that a new · kind of piacenta material at the same early increase in the amount of material recovered from the placenta can be preserved if you stick to the solvent

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Placenta extract a sals, preferably an alkali or alkaline earth, susetst and the ingredients of the placenta extract in a salt complex. The complex thus obtained is in similar to the pharmaceutical effectiveness of the placenta material obtained by the process of the said USA patent.

With the i-lacentamaterial obtained according to the invention, the Osteoarthritis can be treated with success

Recruitment Process

The method of the invention represents a further development of the method the USA patent 2 907 695 for the production of pharmaeutxsch valuable material from the placenta through treatment

the placenta with an acidic solvent, such as an acidic, aqueous alcohol, with only part of the placenta in the solvent is extracted, while the remainder forms the insoluble residue by according to the invention according to the method of the USA patent obtained solvent extract a salt, preferably an inorganic alkali or i ^ rdalkallsals, admixed which forms a pharmaceutically acceptable salt complex with the ingredients of this extract, which in analog The way in which magnesium phosphate works in a complex manner is nus one acidic, aqueous ethanol extract according to the method of the USA patent 2,907,695 is obtained by precipitation with a base.

The placenta extract can be both inorganic and organic Alkali and bradalkali salts added - »earth. A few examples

for this are: / potassium chloride, -gluconate, -bicarbonate -cltrate * oil-based and tribasic potassium phosphate,

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-aalicylate aowia -lactate, dibaaiaobaa and tribaaiacbaa Sodium phosphate, sodium laotate, salicylate, -bicarboot, -chloride, -citrate nowie -sulphate, dibasiaches and tribaaiacbaa calcium phosphate, calcium gluconate, calcium lactate, carbonate, citrate, levulinate, aandelate, ethionate, -chloride aowia -aalicylate, dibaaiaches and tribaaiachaa Magneau phosphate, magnesia lactate, carbonate, citrate, -aalicylate, -thioaulfate aowia -trisiliopiat, Gtrontiualactat, -citrate, -carbonate aowia -aalicylat, tribaaiachaa Gtrontiuaphosphat

From this, Hioaphatan is the precaution gagaban «The beaten results aind with 2-baaiachen Maganaaiu ^ phoaphat (HgHfO _4. Obtained with daa 5-baaiachan Magnaaiuatphoaphat (Μβ (Ρϋ ₄ ) ₂ .5Η ₂ ⁰ ^ ^{and b # l of the} porenteral Application because of the phamaaeutiach of valuable material in the application area slight indications have been observed. Determinations of the noble simple species which would be obtained when using 2-basic magnesia phoaphates have shown that the erieltesj pharaaeutiacha growth is the same, but less pronounced the weight was about 60 to 70, including 65 parts of the dibasic Maajnasiuaphoephatkoaplexes 15 parts of the magnesia-phosphate coaplex obtained by the method of the US Pat. No. 2 90,695, as disclosed in FIG the substances seem to be the same.

The substances obtained from the invention are called "pharaaeutically contractual SaIBkOaPIeXa ¹ *, sB · daa by Dimagneeiua-

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received phosphate · material is ale "dibasic magnesium phosphate complex", if the origin is to be indicated, or more generally referred to as "Magnesiuaphoephatkoaplex".

Mahr ine individually · going: You finely divided placenta becomes with a suitable solvent, e.g. acidic, aqueous methyl or ethyl alcohol, or any other suitable solvent alt Extraction capabilities for the placenta material in contact brought. Rieh is particularly suitable for this

W of 75 % ethyl alcohol and 25% A'asser, the ait QaIs- or

Sulfuric acid, preferably at an initial pH of about 2.7 · acidified iat After the extraction, which is carried out for about 2 hours at 2o to $ o ^e C, the mixture is left in the cold swecKaässig · 4 to 5 hours at ⁰ C - stand to achieve the precipitation of the proteins su. This leaving it in the cold is not absolutely necessary and can be omitted. The extraction solution is then separated from the residue and the salt, for example an alkali or alkaline earth metal, is added to the solution. But you can also add the SaIs after the precipitation of the ilacentaamateriala in the manner described in US Pat. No. 2,907,695.

The first case is after the extraction solution has been separated off ▼ cm residue and after adding the alkali or alkaline earth salt with a base, such as aaaoniua or alkali hydroxide, the pH value the sunächet acid solution to about 9 to 1o, e.g. 9.6, increased, wob «! Precipitation of the rlacantaa material begins, its insulation in the manner indicated in U.S. Patent 2,907,695 can.

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In the second case, the placenta material is isolated as a magnesium phosphate complex by first - as in the USA patent 2 907 695 stated - the verbose material with a base the acidic Extraktlonelöaung falls and separates and the so obtained Liquid, which also contains further placenta material in solution, an alkali or Krdalkali is added, whereby one besides the placenta material obtained by adding a base receives a salt complex of the placenta material

After the base solution to the acidic extraction solution, the precipitated uagnesium phosphate complex separated and from the Filtra * the solvent can be removed by evaporation. A paste-like extraction residue separates out in water dissolved and mixed with the salt used according to the invention can be, whereby the salt goes into solution, at the same time the reaction medium becomes cloudy and the sals complex of placenta material precipitates · The precipitate can be separated and used for the isolation of the magnesium complex in the USA patent 2 907 695 specified iron be worked up.

Under the term "mixing the solvent-containing placenta extract with a “salt” is both the mixing of the solvent-containing Placenta extract as well as the extract that is after Removal of the solvent converted into a new solution has been to understand with a GaIs

The amount of salt is 10 to 5000, preferably 100 to 3000%, on the weight of the deposited in the evaporation of the solvent uxtractionary residue related, oil-based magnesium

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phosphate or di-equivalent ». Meng · other salsa · at deei the first-mentioned procedure, i.e. if there is an acidic solvent extract xugesetst, the salsaenge, i.e. the henge that is used when the other procedure follows, can . correspond to the specified quantities.

In the process, the invention is based on temperature and pressure not critical. The characteristic of the process according to the invention is merely the addition of the salt used according to the invention su dea solvent-containing placenta extract, in which «das Basically be soluble what.

While according to the US patent 2 907 695 process Starting material only used human placenta wxrd, can common invention gen * generally the rIacenta of mammals, e.g. cows. The cow piacenta becomes pharaaseutic valuable Piacentaamaterial obtained in good yield and can be used in hua therapy

example 1

1 kg of the adjacent placenta is kept at 2o to> * C for 2 hours aqueous ethyl alcohol (75% alcohol), which has a pH of 2.7, brought into intimate contact, the mixture obtained was left to stand in the cold, filtered and the piltrate - like la BLspiel of UBA patent specification 2 907 693 stated - to pH 9 «6 set «The precipitate, probably a magnesium phosphate complex, is separated off and the solution obtained, which still contains further extract portions, which are not precipitated with the base

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will be »ur extraction of suaätsliohea flacenta material subjected to further work-up.

For this purpose, the solution obtained is placed on a water bath evaporated from 100 * C but dryness, whereby 20 g of a dark brown sticky Extraktionarilckatende ■ are obtained. 100 g of the rüokatandea are added in 227 g of line wafers 5 ^ »5 * C, which quickly becomes a dark brown solution This forms a pH value of 4.5. This solution is β and Bt 454 g water and 500 g dibaaiachea magnesium phosphate su and stir it well for about 1 hour at room temperature. Ea sets'

Foam indicating the reaction. Then add 9 * 10 g Water of 5 *% 5 * C and another 500 g dibaaiachea magnesium phosphate see below, continue stirring for about an hour and then leave the resulting solution stand for three hours at room temperature · The pH of the solution is 7 * 0 · The solution is then poured into a Pyrex evaporation aachale and feeds in an oven 70 ° C for three days by evaporation. You get a hard, Ate-like, caramel-colored product, daa in a mortar rubbed into the pharmaceutically valuable placenta material can be·

Examples 2-13

Per 100 g of the brown obtained by the process of Example 1 Extracta were in 500 ecm of water at 66 * C see below dissolved in a clear solution, then 1 kg each below specified alkaline earth or alkaline salts sugesetst (10 wt per weight of dissolved substance) and everything thoroughly

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mixes. The mixture obtained was read within 3 days evaporate to the back at a temperature of 55 ° C, whereby the pharmaceutically valuable salt complex was formed. That Froduct was a hard, etein-like material that then ground to a powder in a mortar.

Table 1 Example no. 2 *
Caloium gluconate 3 Calciualactat 4th dibasic calcium phoephate 5 tribasic calcium phosphate 6th dibasic * magnesium phosphate 7th tribasic magnesium phosphate 6th Potassium chloride • 9 Potassium gluconate 1o dibasic sodium phosphate 11 Sodium lactate 12th Sodium salicy1ate 13th Strontium lactate

For the preparation of a form of the placenta material that is suitable for parenteral administration, 1 to 3 * e.g. 2 g of the powdery material obtained are mixed in 3o ecm of a sterile normal saline solution (0.9 Hael), which preferably contains 0.5% benzyl alcohol and 0.5% as a preservative. contains 01% Butaben, dissolved by heating on a water bath at 100 ⁰ C for 3o minutes · foams shows the reaction during this time · the resulting solution is cloudy and has a pH of 7 »0th

this material is suitable for parenteral application, e.g. intramuscular application in patients with rheumatic disease

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Arthritis, osteoarthritis or intermittent claudication «. The powder made by grinding in a mortar can also be taken orally, e.g. in capsules.

Example 14

Placenta was established using the US Pat. No. 2 90? 695 brought into contact with aqueous ethyl alcohol at about room temperature, then left to stand in the cold, bring the protein material to precipitate and the clear solution obtained after removal of the protein material evaporated to dryness, whereby the acidic material was obtained as a brown residue. 100 g of this brown residue 1 / Vaaaer of 140 ° C was dissolved in 1 dissolved, the solution 1 was added kg dibasiaches magnesium phosphate, stirred for Mirchmischung and thereafter evaporated in an oven at 70 ⁰ C to dryness · The dry Fostteilchen were removed from the oven, a 75UgO solution made of normal saline sterigier of dry matter, the then 45 limit has been s at 100 ⁰ C in water sterilized ·

Example 15

It became a 7% saline solution of the placenta-eel coaplex of the example 14 prepared and the solution after sterilization of a Group of patients with osteoarthritis symptoms in doses of 2 ecm injected

The treated patients showed a substantial improvement regarding osteoarthritis

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Example 16

Of 84 patients suffering mainly from osteoarthritis of the knee Suffered, no intra-articulars six months before the start of the experiment. Had received steroid, phenylbutazone, or Aralen treatment and put on a basic program of rest and aspirin 42 patients were given 1 cc of a solution of a placebo and 42 patients received the 7% saline solution twice a week Example 15 parenterally injected · Mach 6 »times the Injections reduced to one per week.

Complete bone and general examinations, chest x-rays, electrocardiographs, the Bucket test, blood count, sedimentation velocity, urine test, Radiological examinations were carried out and both patient groups were observed for one year.

The trial showed that of the 42 patients who received the 2lacebo bekoBuaen had only 15 / »satisfactory response while from the 42 patients who inherited the placenta extract of the invention 91% showed an old animal to very good improvement.

Example 17 In the treatment of steoarthritis of 82 patients with one

the alkaline Salzkoapositionen of Examples 2 to 13 below

The following results were obtained using the double blind technique

obtain:

The pulverföxvigen üalzkoaplexe of Examples 2 to 13 were 5% solutions transferred, while each 1.5 g of the obtained

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Alkali and bradalkali salt complexes in 3o oca sterile normal Brine solution dissolved in a simmering water bath for 35 minutes sterilized at 100 ° C and then cooled to room temperature lieaa.

The treatment dose, the 4a parenterally up to twice a week appÜEiert was 2 to 5 cc «. After an improvement occurs the dose in osteoarthritis was at intervals of 2 to Weeks reduced.

The response to the placebo was 2o to 25% in all! the reaction to the group of alkali salts So to 89% «whereby the effectiveness of the composition of the invention in treatment of Oflteoarthritia becomes evident.

Has further old oral therapy in Üsteoarthritis achieved very good result · Oer aalskomplex was in a einsigen daily dose of 0.6 to "ewandt · Here an excellent response of 70% and a good response in 15% of cases was achieved · With dea irlacebo (a similar-looking capsule with milk sugar) a reaction of 18 am 24 -r was achieved

Tolerance symptoms and side effects did not occur in one only case of those treated with the ski compositions of the invention Patient on.

All the patients treated in these oeries suffered from different ones Types of osteoarthritis involving the knees, hands, neck, hips and shoulders in most cases »All had

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severe pain, swelling, and difficulty walking or sleeping. Neither was bit one of the previous ones Treatments, including cortisone, - ^ utasolidin, diet mineral water therapy, Bee venom, gold, etc., have been given greater relief. All actually had one certain disability. Usually your treatment was both oral and Dei Injections gave a similar and very satisfactory response received · The pain and swelling disappeared and an improved sense of well-being set in · The patients were able to her «put away crutches and sticks and live normally without opiates ·

Chemical identification of the placenta material obtained: Analysis of the procedure according to the U8A patent 2 907 received material!

One by precipitation from the acidic aqueous ethanol solution The sample obtained has a soft, waxy consistency after drying on the steam bath. Alien »never has one ammoniacal odor in alert state after removing dea steam bath · mach the infrared absorption curve appears the compound is a primary amine, possibly of the -CH-HHg type (aliphatic), presumably to be an amide su. At 6.3 and 7 »1 μ are gangs. These can be due to aaino acid bonds, possibly also attributable to leptide bonds leg. Another possible explanation for the band at 6.3 μ is that the compound is a carboxylic acid (zwitterion)

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In the chemical analysis, the following species were obtained

magnesium 17.4 % (nothing) phosphate 47.1 % Total ticketoff 5.7 % ammonia organic nitrogen,
calculated from the difference O, 6% sulfur 0.0 % urahydrogen 5.95 % Water (as moisture) 13.5% organic fiber,
calculated also the difference o, 3%

The organic part of the beet, 3.6% of the total, had the following composition:

Carbon (analysis) 1.79%

.Vase only off "0.3 %

Nitrogen "0.6% Oxygen (difference) ο, 9% The ash content of the -frobe was 49.63% According to these chemical and organic analysis data and dea So the sample seems to have the following compositions:

Mg ₃ (P0 _{4) 2} · 8H ₂ 0 76.8% i

H ₂ O 13.5 %

NH ₃ 6.2 %

organic components 3.6%

YJB was a book on traces found · The analysis data of the organic part of the J. robe rode * u an empirical • "Ormel ^c of about 4 ^ ^H 28 ^ 4 ^t) 5 ° & ^{he inr"} "multiples ·

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It should also be pointed out that the flacenta materials of the invention, they will old various other cheaipchen Combined compounds, ways of treating different Diseases offer. For example, the brown extract residue of Example 1 with 25 wt specified materials are mixed. With the mixtures obtained could if they were in the form of 0.3 g capsules three times

! administered daily, improvement in rheumatic arthritis _ thritis can be achieved.

A) Chemicals of the salicylate group, such as sodium salicylate, acetal

salicic acids,
BO compounds of the p-aminophenol type, such as Acetamilid and Acetp-

phenetidine
C) Pyrasolone derivatives such as antipyrine and aoinopyrine.

Furthermore: the brown residue of Example 1 is brought into solution, it can be combined with numerous sulfonamides, when the sulfonamides are added to the solution and the solvent is evaporated and forms a dry powder · £ s has been found that with a dose of 0.3 g at intervals of 3 hours a good clinical response has been achieved in rheumatoid arthritis patients, some examples of the sulfonamides are: SuIfameraxin, SuIfacetamid, Sulfadiozin, iSulfamethazin, SuIfapyridin, Sulfadimethoxine.

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Claims (2)

I 4b / 908 godparents * 'Tnepriicho 1) Process for obtaining plabeat material by extraction the placenta ait «Inen eauren LOanngaaittel and, if necessary after previous removal of the proteinsPrecipitation of the placenta * nateriale in the alkali-low area, dadaroh marked, deea en · Plaoentaeztrakt before or after the low-alkali precipitation inorganiciaohea or organieohea alkali or alkaline earth as sugar acetate will. 2) Method according to claim 1 «thereby gekennseichnet ,. daae out the piltrate obtained after the low-alkali precipitation as a solvent drained, the received · Büokatand in laaaer gelöat and with an inorganic or organic alkali or Rrdalkali as Toraetst will BAD OBiGiNAL 009820/15 «!