CS197095B1 - Process for preparing antibiotic citrine from microorganism penicillium janthinellum biourge - Google Patents

Description

SUMMARY OF THE INVENTION The present invention provides a process for the preparation of an antibiotic citrinin from a novel production microorganism Penicillium janthinollum Biourge.

Biologically active secondary metabolites of microorganisms include antibiotics, alkaloids, mycotoxins, enzyme inhibitors, growth promoters and the like. Citrinin is a metabolite of several microscopic fungi from the genus Penicillium, and its production by Penicillium janthinellum is not reported in antibiotic producers (eg J. S. Glasbyt Encyolopaedio of Antibiotics. John Wiley and Sons, London, 1976, pp. 134-135). Its antibacterial properties have been known for a long time, but recently its antifungal and insect activity has been recognized.

The present invention contributes to addressing the need for novel biologically active compounds and provides a process for preparing citrinin in a crystalline state,

Process for preparing citrinin from Penicillium jenthinellum S-52 deposited under CCM number P-391 in Ss. collection of microorganisms of J.E.Purkyně University in Brno, Tř. The invention is based on the principle of cultivating the microorganism Penicillium janthinellum S-52 in a liquid nutrient solution containing carbon sources in the form of carbohydrates, organically bound nitrogen and minerals under temperature conditions of 24 to 30 ° C. and at a slightly acidic pH, the filtrate was fermen197 095

197 095 of the reaction medium or the precipitate obtained by acidifying the filtrate with hydrochloric acid to a pH of up to 2 is extracted with an organic reagent of limited miscibility and water, preferably chloroform or ootonic acid esters, dissolved after evaporation in methanol, acetone or benzene and crystallized to obtain crystalline oitrinin ·

In the following, the invention is explained in more detail by way of examples of procedures without being limited to these.

Example 1

Promoted culture of Penioilllum janthinellum S-52 on wort agar in 1000 ml Roux culture flask is poured with 40 ml sterile water and spores are released, 10 ml spore suspension is added to 1000 ml culture flask with 150 ml sterile nutrient solution of the following composition: drinking water 1000 ml, glucose 45 g, saoharose 45 g, sodium nitrate 2 g, secondary potassium phosphate 1 g, magnesium sulfate 0,5 g, potassium chloride 0,5 g, ferrous sulphate 0,01 g, concentrated corn extract (50% dry matter) 10 g, pH adjusted to 6.5 before sterilization. It is cultivated on a rotary shaker with 220 rpm ** ¹ and a circle radius of 5 µm at 38 ° C for 24 to 30 hours. ml culture vessels. It is cultivated on a rotary shaker for 5 to 6 days under the above conditions, when maximum oitrinin production is reached.

Example 2

The preparation of the spore suspension and the vegetative inoculum of Penicillium janthinellum S-52 are the same as in Example 1. The fermentation conditions are the same but in a nutrient solution of the following composition: drinking water 1000 ml, sucrose 40 g, concentrated corn extract (50% dry matter) g, secondary potassium phosphate 4 g, the pH is not adjusted prior to sterilization. Cultivation takes 10 to 11 days to achieve maximum oitrinin production. In this case, the production of the antibiotic is about three times that of the nutrient solution of Example 1,

Example 3

The myelium of the Penioilllum janthinellum culture after the fermentation of Example 1 is complete or is separated from the culture solution by filtration and the filtrate is filtered through a pad of diatomaceous earth. A third volume of chloroform was added to 5 L of the filtrate and the pH was lowered to 1.5 by addition of hydrochloric acid before the two phases were mixed. The strips were thoroughly mixed with a stirrer and then separated. The chloroform extraction was repeated two more times. The combined chloroform extracts e are freed from water by filtration through a layer of anhydrous sodium sulfate and the filtrate is evaporated to dryness. 5 g of crude oitrinin preparation are obtained from the culture of Example 2.

Example 4 g of the crude preparation obtained according to example 1 are dissolved in 50 ml of carbon tetrachloride, the solution is filtered and thoroughly mixed with 20 ml of ethylene glycol to which they pass

197 095 accompanying brown pigments. After phase separation, the yellow-fluorescent solution of citrinine in tetrahydrofuran was evaporated to dryness, the residue was dissolved in acetone and crystallized. 500 mg of crystalline citrinin is obtained.

Claims (1)

Process for the preparation of the antibiotic citrinin from Penicillium janthinellum Biourge, characterized in that Penicillium Jenthinellum S-52 CCM P-391 is cultivated at temperatures of 24 to 30 under submerged conditions in a liquid nutrient solution containing carbon sources in the form of carbohydrates, organically bound nitrogen and minerals. ° C and at a slightly acidic pH, the fermentation broth filtrate, or the precipitate obtained by acidifying the filtrate with hydrochloric acid to a pH of 1 to 2, is extracted with an organic reagent limited to water, preferably chloroform or oic acid esters, dissolved after evaporation in methanol, acetone or benzene and undergo crystallization.