DE1810033A1 - Process for the production of deoxycholic acid - Google Patents

Description

Dr. Ing.Jürgen Schmidt Dr. rer. ηεί. Bruno Reiizner

Patcr.iänv / Lltä

8 Munich 2, Hsrmann-Sack-Str. 2

20th November 1968

Regarding patent application

METHOD OF MANUFACTURING 7ON DESOJOfCHOLSAURE

The invention relates to a new method of manufacture of extremely pure deeoxycholic acid, which can be used for bacteriological purposes.

The currently commercially available types of deaoxyoholaic acid and are mostly not suitable for use in bacteriological Fertile soil. The deaoxycholic acid which can be used for such purposes must have special requirements with regard to the

old.

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Fulfill purity and must first and foremost be free from fatty acids. ¹

Deoxycholic acid (according to the chemical designation 3cC "l2 * x. -Dihydroxycholanic acid) forms adduct compounds, so-called quholeinates, with numerous different substances, including fatty acids, and similar compounds with aromatic and heterocyclic compounds, such as benzene, morpholene, etc. Chol-einaäure also consists of such fatty acid choleinates »In the industrial preparation of this product from binder bile, the alkaline hydrolysis also releases the fatty acids from the raw material and these immediately form the above-mentioned adduct compounds with the deoxycholic acid formed *

But since such fatty acid choleinates are used for bacteriological purposes »Mui from these dl® free deoxy alcoholic acid © after removing the fatty acids cannot be used. to be isolated «,

In the previous literature on the purification of olealic acids and the production of deoxycholic acid, this important circumstance was neglected until the last few years, and the literature often made no distinction between free deaoxycholic acid and the choleinates mentioned above containing adduct compounds . For example, after L * 0. Paterson (B. Ρ «695 504 ₉ 1953} separates deoxycholic acid from an aqueous solution of crude bile acids at pH = 6 with morpholine from the cholic acid which is soluble in morpholine at this pH value.

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SDle obtained in this way as an undissolved residue But deoxyoholeic acid is not the essential free compound, but their adduct formed with fatty acids » Kaoh R.H. Sifftrd (USP. 2,606,912) becomes deaoxycholic acid with the help of benzene, j is separated in this way the deoxycholic acid separates in the form of a ben- ■ ol adduct, thus also in the form of a cholic acid and is purified by recrystallization. From the In this way purified gholeinic acid can but the free deoxy alcoholic acid only through complicated further Steps released with very poor yields and dated Bemol to be insulated.

In contrast, the pure, fatty acid-free Deoxyoholeic acid according to the present invention by a simple, single step process obtained directly from the fatty acid-containing choleinates obtained by processing the raw bile will.

The invention stems from the discovery that the fatty acid adduct compounds containing about 7 # fatty acids the deaoxycholic acid are set and the fatty acids can be selectively deposited »if one e.g. Fatty acid cholelates dissolved in methanol are mixed with urea. The urea forms solid complex compounds with the fatty acids, which from the solution in crystalline Form fail »while the one released in this way Under such circumstances, deoxycholic acid does not form an adduct with the urea, but in its pure state

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remains in the solution. The complex compound of the fatty acids formed with the urea is, according to our experience, a much more stable product, 'rock the product compound of the fatty acids formed with the deso cfaolsäfcre' and so when the fatty acid ahoieinafr® is reacted with urea these aereetfist 'and ύ, Ιύ fatty acids can be in 'The insoluble crystalline urea complex is separated from the deoxycoholeture *, which means that ¹ EUe of the solution can be obtained with high * about 75 to 80% yields * compared to the higher yields of 25 to 30% of the well-known Terfatoen © inen means considerable progress *

The object of the Srfladung is therefore a Yerfahreit äsur producing relner _f fettaäurefreier "SBU bakterioj loglsohen purposes suitably Desoxjoholsäure aöö Fettsäureoholelnaten" which thereby Lat gekennsieiohnet "daf to the Fettsäurecholeinate In organic» treated sviöokraKßig in methanollscher Löeung with urea "the SiOH depositing in solid form the fatty acid-sweetener complex is separated off and the deoxycholic acid or its alkali metal or alkaline earth metal salt is isolated in known catfish from the solution.

To form the fatty acid-urea Somplexes the urea is suitably, in solid, crystalline form of the ssu added ait also a urea organic solvent dissolving _t advantageous with methanol solution of the OhOleinsäureη produced; the formation of the complex is achieved by heating the mixture _t, expediently by boiling it under

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ff

Reflux promoted, then the ReaktionsgemlBch is cooled, advantageously to about 5 ⁰ G, and precipitated in crystalline form fatty acid-urea fComplex is separated by filtration or centrifugation "For the sake of purity of the product obtained it is advantageous to" simultaneously with the urea and Add activated charcoal to the solution of choleic acids.

After removing the fatty acid-urea complex becomes the deoxycholic acid from the remaining organic Solution in a manner known per se, e.g. by evaporating the solution, dissolving the residue in aqueous sodium hydroxide solution and precipitating the deoxycholeic acid by the addition of sulfuric acid, isolated in pure form. Do you want that Deoxycholic acid is obtained in the form of an alkali salt, then after dissolving the evaporation residue in alkali the alkali salt solution obtained is not decomposed by the addition of acid, but immediately to dryness evaporates.

The deoxycholic acid obtained according to the invention is sufficient Immediately, without further cleaning, completely meets the quality requirements of bacteriological application. That is from the gelation ability of the product according to the invention visible! the DeBOxycholic acid obtained according to the invention forms in 1% aqueous solution at pH = 6.8-7.0 37 0 a crystal clear gel in 24 hours. The choleic acids, which contain the deoxycholic acid in the form of an adduct compound show no such ability to form gel.

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The method of the present invention is illustrated by the following Examples illustrated in more detail »

example 1

50 g of olelic acid (mp 187 ° 0) are dissolved in 1000 ml of methanol. The solution is mixed with 200 g of urea and 0.5 g of activated charcoal and heated to the boil while stirring and then the solution containing activated charcoal is boiled for 2 hours with stirring. The mixture is then ^{cooled to +5 0} C and after 8 hours of standing the precipitated crystalline carbonaceous urea is filtered off. The crystals are washed with 100 ml of methanol cooled to 5 ° C and sutured to dryness. The carbon-containing crystals of the fatty acid HBternatoff-Xoinplexes are discarded

■ "The methanolic - solution is evaporated to dryness in the TakMum» the solid residue is dissolved in 400 ml of distilled water * with the addition of sodium hydroxide solution at pH - 9 and filtered until clear. - The solution is treated with 10% pure sulfuric acid to pH - 2 acidified »whereby dl® free besoxyclxolic acid is precipitated. The mass obtained on tile white is first washed with tap water, then with distilled water until it is completely freed from urea or» sulphate ions »

After drying, 40-42 g of fatty acid-free, pure free deoxycholic acid are obtained j F. 172 ⁰ C $ [ex.] ² ? * + 55 ° (c = 1, in 96% ethanol).

Example 2

In an enamelled autoclave with a volume of 250 liters

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r »

. content «earth IO kg of oil alloys in 200 liters of methanol.

\ solves * Oils fctSBUng is made with 35 kg of urea and 150 g of JUctlv-

j ■ ■ ·· ■ -.- i --- '■ · r ■ ■, ■

! kohle Vjtreetet »Dag'Mix is boiled while stirring obtained and stirred under reflufl for 120 minutes. The suspension is then cooled by water cooling Huflere to 25 ° C and discharged provided in a cooling jacket and stirrer old Xrlstaiiiilergefäa of a 250 liter capacity "paa mixture is cooled +5 ⁰ C under Rühren.auf and held for 8 hours at this temperature. The crystal mass is then put on a centrifuge and 'rooofed', 'discharged'

i) le crystals are washed with 20 liters of methanol cooled to 5 ° 0 «the washing liquid with the The mother liquor is combined and the carbonaceous urea crystals which have been defused are discarded. The united The methanol phase is pumped back into the enamelled 250 liter autoclave that was initially used and sealed in vacuo,

The dry residue left in the autoclave is dissolved in 200 liters of distilled water with the addition of 800 g Pure Tatriuahydroxld dissolved and the solution, which has a pH value of about 9, is filtered off through a lens filter.

The pure solution obtained is acidified to pH = 2.5 with 10% pure sulfuric acid while stirring.

After stirring for 30 minutes, the precipitate obtained is separated off with the aid of a filter centrifuge. The to- * traces of urea and sulphate left behind are caused by

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Wash first with tap water, then completely removed with distilled water

The moist leather cover obtained is suspended in an enamelled 250 liter autoclave in 80 liters of distilled water and dissolved by adding 10% pure latronlsjig © to pH 8 _a 2 to 8 5 200 to 220 ⁰ O entrance and at most 16O ⁰ G exits temperature dried "Bs are 7» 5 to 7 »8 kg of pure product preservation th, of which at pH * 6.8 to 7.0, a crystal-clear QEi

20th
$ [oc] _D - + 52 ° (c »1, in 96 # ethanol) is formed.

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Claims (3)

PATBNTAHSPRÜCHEi 1. Process for the production of ^ pure, fatty acid-free Deoxycholic acid from fatty acid choleinates »thereby characterized »that the fatty acid choleinates dissolved in organic solvents, expediently in methanol Treated with urea, separates the precipitated fatty acid-urea complex and from the Solution the deoxycholic acid or its alkali metal or Alkaline earth metal salts isolated in known catfish. 2. Yerfahren according to claim 1, characterized in that the alcoholic, expedient methanolic solution of fatty acid cholelates mixed with solid urea »the mixture is heated and after Cooling down, the solid fractions are expediently removed by filtration or centrifugation. 3. The method according to claim 1 or 2, characterized in that one in the formation of the fatty acid-urea complex also add activated carbon to the reaction mixture. - 9 S09828 / 1623