SU122846A3 - The method of obtaining spiramycin - Google Patents

Description

Society of Chemical Plants Rhona-Poulenc (France) Actual Inventors Foreigners Nine-m-Varia

METHOD OF OBTAINING SPIRAMICINE

20 nun IS58 were filed for jY ° 60244С / 31 to the Nzobrotesh and

discoveries at the Council of Ministers of the USSR Published in "Bulletin of Inventions L 19 193

It is known that the antibiotic spiramycin consists of three components I, II, P1, which have very similar properties, and is obtained by the naked fermentation of Streptomyces ambofaciens.

A method is proposed that allows to produce less toxic variants of spiramycin (II and III) by suppressing the formation of a more toxic variant (spiramycin I). This is achieved by adding acetylating or propionating radicals to the medium. Example.

260. Or the following medium is introduced into an Erlenmeyer two-liter flask:

Corn infusion (50% dry extract) 40 g Glucose20

Sodium Chloride5

Magnesium sulphate

Plumbing water 1000 ml

The pH is adjusted to 6.8 with sodium hydroxide and then more:

Calcium carbonate 5 g

Soybean oil4 ml

This medium is sterilized for 45 minutes. at 120 After cooling, it is seeded with a strain of Streptomyces ambofaciens (NRRL 2420 in the galaxy). Then the culture is stirred on a tr bitch for 48 hours.

At the same time, 5 ml of the following medium was added to 300 ml Erlenmeyer flasks:

For the visitor

Corn infusion (50% dry extract) 35 g Glucose50

Sodium Chloride20

Primary potassium phosphate2

Magnesium sulphate1

Plumbing water 1000 ml

The pH is adjusted to 6.8 with the addition of caustic soda, followed by the addition of:

Calcium carbonate 5 g

Then propopamide is administered in the amounts indicated in the table below, giving it1, the results of the experiments performed.

Erlenmeyer flasks sterilized for 20 minutes. at 120, 4 ml of culture from two-liter flasks are cooled and seeded, and then shaken on a truncheon at 25. Analyzes are carried out on 6-ii, 7th and 8th days in order to determine; relative outcomes of three spiramycins.

PRI me R 2.

Into a two liter Erlenmeyer flask, 250 are introduced, or the following medium:

Corn infusion (50% dry extract) 4-0 s Starch20

Sodium Chloride5

Magnesium sulphate1

Primary potassium phosphate2

Tap water up to 1000 ml

The pH is adjusted to 6.8 with the addition of caustic soda, after which it is introduced: Sodium carbonate 5 g

This medium is sterilized for 45 minutes. 120. After chilling, they are seeded with S. arnboiaciens culture and shaken on a bitch for 48 hours. at 25 °.

At the same time in Erlenme 1, the flasks for 300.

Corn paste (50% dry extract) 45 g Glucose5

Calcium carbonate25

Tap water to a total volume of 1,000.

The pH is adjusted to 7.0 with the addition of caustic soda.

After that, the medium is sterilized for 30 minutes. at 120 p.p.yu cooling each erleimener flask is inoculated 4 ml previously on: a refill culture. The mixture is then shaken on a tr knot at 25 °. After 48 hours fungus development is very satisfactory. Thereafter, 20 ml of a sterile EOI solution containing 12 g / l of pure spiramycin I in the form of osigone and 6 c1 / l of sodium propionate are added to each flask. The culture is shaken for 24 hours, then the contents of all Erlenmeyer flasks are mixed.

Chromatographic analysis of the broth culture shows that it contains 2.3 g / l of unstained syiramycin I and 1.4; mixture consisting of pz 20u spiramycin P and. spiramycin III.

PRI me R 3.

In erlenmeyero; -58C1 e 300 ml flasks are introduced by about 40.,;.; sochuyuyuG; Wednesday:

Ammonium chlorides6 s

Glucose25

Calcium carbonate20

Tap water to a total volume of 1000 ml.

Sterilize, inoculate, and culture the fungus under the conditions described in Example 5. After 48 hours. An aqueous sterile solution is added in 20 g each with a content of 9 g / l pure spramine 1 as a base and 21 g / l propionamide.

After three days of incubation and chromatographic analysis, a conversion of 1.86 g / l of spiramycin I to a mixture of 55% spiramycin II and 45% spiramycin P1 is detected. 1.14 g / l of the scroll. Mycin I remains unchanged.

At measure 4.

A culture of S. ambofaciens is prepared under the conditions described in Example 3. The spiramycin solution of base I contains another 6 g / l sodium propionate. In this case, chromatographic analysis reveals the conversion of 2.04 g / l of syiramycin I into a mixture of 11% spiralization II and 89% of spiramycin P1; 0.96 g / .g of the starting spnramationa remains unchanged.

Example 5

In a 300 ml Erlen. Meier flask, 50 ml are introduced, followed by the medium:

Ammonium nitrate13 s

Glucose50

Calcium carbonate25

Tap water to a total volume of 1000 ml.

Flasks are sterilized with 30 .mip. at 120, cool and seeded with 5 MI cultures, the preparation of KOTopoii is described in Example 2. After two days of incubation at 25 ° with shaking on a fork, each flask} is poured with 5 ml of an aqueous solution containing 100 g / l of pure spiral I bases, 38 g / l of propioic acid and 10 g / l of cobalt trihydrate. The pH of this solution is pre-adjusted to 6.5.

Eh 122816

Law 122846-4 by adding caustic alkali. After a three-day incubation, analyze by paper chromatography. 8.70 g / l spiramycin I converted; yield 9% spiraminin P and 91% epiramycin III.

Example

In a 300 ml Erlenmeyer flask, pour 50 ml of the following medium:

Ammonium Chloride 10 g

Glucose50

Calcium carbonate25

Tap water to a total volume of 1000 ll

The flask is sterilized, seeded and incubated under the conditions described above.

After 72 hours the culture is centrifuged and the liquid is decanted. The precipitate is re-inoculated in 50 ml of pure buffer solution, contained in a 300 ml Erlenmeyer flask. The buffer contains 9.85 g / l of glacial acetic acid and 4.9 g / l of sodium acetate and has a pH of 4. In addition, the buffer also contains 6.45 g / l of epiramycin I hydrochloride and 1 g / l of cobalt hydrochloride . A suspension of this mycelium is incubated for 80 hours. at 25 ° with shaking on a cable and chromatographed on paper. As a result, 2.40 g / l of epiramycin I is converted into a mixture of the remaining spiramycins (II and P1) in relative sizes of 90:10.

Subject invention

The method of producing epiramycin, carried out by fermenting Streptomyces ambofaciens, characterized in that, in order to obtain less toxic variants of epiramycin (II and III), acetylating or propionating radicals are added to the medium.