DE1213844B - Process for obtaining pure vitamin B. - Google Patents

Description

FEDERAL REPUBLIC OF GERMANY

GERMAN

PATENT OFFICE

EDITORIAL

Int. α .:

C07d

German class: 12 ρ-10/05

Number: 1213 844

File number: P 28466 IV d / 12 ρ

Filing date: December 22, 1961

Opening day: April 7, 1966

The invention relates to a process for obtaining pure vitamin B ₁₂ from contaminated solutions by precipitation with polyhydric phenols with subsequent separation and dissolution of the precipitate thus produced, which consists in adding about 20 times the amount _{to the neutral aqueous vitamin B 12 solution} of the vitamin B ₁₂ contained therein is added to tannin, the separated precipitate is dissolved in dilute acetic acid and the vitamin B _{12 is obtained} in a known manner from this solution, if necessary after carrying out further cleaning measures.

Vitamin B _{12 has} already been obtained from its aqueous solutions by precipitating it as a vitamin B ₁₂ phenol complex and breaking this complex down into its components. The known method, in which a mono-, di- or triphenol was added _{to precipitate vitamin B 12 , could only be applied to vitamin B 12} solutions which contained at least 0.5% vitamin B ₁₂ . The method according to the invention is not subject to any such restriction and can be applied to solutions with a very low vitamin B _{12 content.} It makes the use of large amounts of solvents superfluous, which were previously _{required for the production of vitamin B 12} from cell suspensions and fermentation solutions with a low vitamin B ₁₂ content, and thus brings about a significant simplification in terms of plant and labor, time and effort As far as costs are concerned.

To carry out the process according to the invention, it is expedient to start from a prepurified vitamin B ₁₂ solution, which was obtained in the following manner:

A suspension of _{microorganism cells containing vitamin B 12} is heated to 100 to 120 ^° C. for 10 minutes and then centrifuged. The cell-free solution containing vitamin B ₁₂ is purified by adding calcium oxide. The addition of calcium oxide causes the impurities to be largely precipitated without impairing _{the solubility of the vitamin B 12 remaining in the solution.} No protection is claimed for the production of the pre-purified vitamin B _{12 solution.}

According to the invention, tannin is added to the solution pre-purified in this way, after it has been neutralized, in excess of the vitamin B ₁₂ contained therein. The precipitated vitamin B ₁₂ tannate is centrifuged or filtered off and dissolved in dilute acetic acid. The total solids of this solution is about 0.5% and the vitamin B ₁₂ content is about 0.010 to 0.015%. The in-solution process for obtaining pure
Vitamin B ₁

- "12

Applicant:

Pierrel S. p. A., Milan (Italy)

Representative:

Dr. W. Beil, A. Hoeppener and Dr. H. J. Wolff,

Lawyers,

Frankfurt / M.- Höchst, Adelonstr. 58

Named as inventor:

Dr. Ivan Rolovich, Naples;

Dr. Luciano Devescovi, Capua, Caserta;

Giuseppe Pellone, Portici, Naples (Italy)

Claimed priority:

Great Britain 23 December 1960 (44 301)

The vitamin B _{12 contained} is then purified in the usual way. It is obtained in a degree of purity of more than 95%. Since the volume of the solution obtained in this way is relatively small and its degree of purity is very high, small operating systems and only one further purification by ion exchange or chromatography are sufficient to obtain crystalline vitamin B ₁₂ of high purity. The following example explains the method according to the invention.

example

To produce the starting solution for the process according to the invention, 101 of a cell suspension containing about 245 mg of vitamin B _{12 is} heated to 100 to 120 ° C. for 10 minutes while stirring.

After cooling while stirring, 101 pure acetone are added. The mixture is stirred for 30 minutes and then centrifuged or filtered to separate the vitamin B _12- enriched extract from the cells. The extract is concentrated to three tenths of its original volume. The enriched, concentrated aqueous solution has a volume of about 61 and a vitamin B ₁₂ content of 38 to 40 μg / ml, corresponding to a total content of about 235 mg. 1.5%, ie about 90g calcium oxide are added to it while stirring. The pH of the solution rises from 6 to about 10. The solution is stirred for 30 minutes

609 557/314

and then filtered. The residue is washed twice with one tenth of the original volume of demineralized water each time. Thereafter, the solution has a volume of about 7.21 and a volume of about 7.21. The vitamin B ₁₂ can be precipitated or crystallized from this solution directly, but the solution is preferably purified beforehand by chromatography on xon earth.

Vitamin B ₁₂ content of about 220 mg or 5. 185 mg of crystalline vitamin B _{12 is obtained} , which is 30 y / ml. The yield is more than 90%. The pH of the solution is then adjusted to 7 by adding normal sulfuric acid.

4.5 g of tannin, ie about twenty times the amount of vitamin B ₁₂ , are then added to this pre-cleaned solution while stirring, after which the mixture is stirred for about 60 minutes at room temperature. A precipitate of vitamin B ₁₂ tannate is formed, which is centrifuged or filtered off. The moist precipitate, which contains practically all of the vitamin B ₁₂ present in the solution, weighs 20 to 25 g. It is slowly dissolved in 500 cm ^{3 of} a 5% acetic acid solution. It dissolves completely. The clear solution has the color characteristic of _{vitamin B 12.} The acetic acid solution of the vitamin B ₁₂ tannate is concentrated to half its original volume, ie to 250 cm ³ , in order to remove the excess acetic acid. Thereafter, the concentrated solution contains 700 to 800y / cm ³ Vita because of its extinction (EJ * _n ) has a degree of purity of 95%.

Claims (1)

Claim: Process for obtaining pure vitamin B ₁₂ from contaminated solutions by precipitation with polyhydric phenols with subsequent separation and dissolution of the precipitate thus produced, characterized in that about 20 times the amount of vitamin B _{12 contained therein is added to the neutral aqueous vitamin B 12 solution} of tannin is added, the separated precipitate is dissolved in dilute acetic acid and the vitamin B _{12 is obtained} in a known manner from this solution, if necessary after carrying out further cleaning measures. The yield is 90% and the Vitamm-B ₁₂ content of the solution is about 195 mg. The overall yield from the original cell suspension is about 80%. Considered publications:
German Patent Nos. 947 193, 842 527;
German exploratory documents No. 1060 549,
012, 1068 429, 1073 151.