CS203352B1 - Method of treatment of fermentation substrate - Google Patents

Description

(54) Treatment method farmentačnoj pody ·

The invention relates to a process for treating fermentation broth prior to isolation of cephalosporin C on synthetic resins.

The recovery of cephalosporin C by purification of the fermentation broth follows its fermentation preparation using microorganisms of the genus Cephalosporium, e.g. strain Cephalosporium acremonium ATCC 14 553, wherein the medium contains a biologically active substance / Crawford K. et al .: Antibiotics production by a species of Cephalosporium. J. Gen. Microbiol. £ (1952), p. 47-59; Hewitt W.

L .: The Cephalosporins. The Journal of Infections Piseares 128 (1973). p. 312th

Such fermentation processes are also disclosed e.g. U.S. Pat. 570 460, no. 565 246, no. No. 570,459, in British Pat. 820,422, U.S. Pat. 3,082,155 and in German patent no. These patents also disclose certain processes for recovering and purifying cephalosporin C from a crude fermentation stage.

Methods for purifying cephalosporin C from the fermentation broth via N-derivatives such as N-haloalkanoyl or N-benzoyl, extracted with suitable organic solvents, are described in US Pat. no. No. 3 739 002, no. No. 3,835,129, in U.S. Pat. NSR no. 2 255 973, no. 2,157,693 and in U.S. Pat. USSR number 282 174.

An important group of isolation procedures are synthetic ion exchange methods, e.g. France. pat. 6. 1,353,113, British Pat. no. 1 210 823, no. 1,036,125, respectively. Australian Pat. no. 251 922, 455 753, Sweden. pat. no. 198,707, 215,561, 198,707, 541,579.

Hydrophilic antibiotics, e.g. cephalosporin C, can be obtained from complex suspension fermentation stages containing a large amount of dissolved organic and inorganic compounds using macroporous non-ionic sorbents, e.g. Type Amberlit XAD-2, Ostion SP-1, Diaion HP-20.

Insulations with these sorbents are contemplated e.g. in US Pat. no. 3,725,400, respectively. USSR pat. no. 350 265.

The condition of a successful isolation process on the resins is the assumption that clear b

filtrate, which serves as a spray on the head of the columns. In the purification of cephalosporins C, the pH is adjusted to 2.7 with inorganic acids. thereby causing protein coagulation.

However, after filtration during the adsorption, there is a gradual further coagulation of some soluble substances, mainly proteins, whereby the impure filtrate reduces the capacity of the sorbents and withdraws the further course of the isolation process, including the regeneration of the sorhents.

The above-mentioned drawbacks eliminate the method of treatment of the fermentation broth, which consists in adding to the fermentation broth after the cephalosporin C biosynthesis at a pH of 2.5-4.0 5 to 30 mg / ml urea, filtering and subjecting the pod to further purification . The process may be supplemented by the addition of 0.5-52 anionic detergent.

It is known in the urea that, in some labile non-covalent bonds, it can disrupt the tertiary structure of proteins and thereby cause their denaturation.

This denaturation effect is very needed because, due to the low thermostability of cephalosporin C, the thermoagulation of the raw fermentation broth is not realistic.

The addition of this material results in a reduction in the dissolved protein content of the filtrate obtained, which ultimately improves the adsorption capacity as well as the regeneration of the resins used.

In addition to the cleaning effect, anionic detergents also act as antibacterial agents, which is very important as a prevention of possible microbial contamination of column fillings.

In this group of detergents, a large group of additives are made up, the major part being sodium lauryl sulfate. TERGITOL, in particular, has proven commercially.

In the following, the invention is explained in more detail in the examples of procedures without being limited thereto.

Example 1

The crude fermentation platform is adjusted with 10 mg of urea per ml of soil, stirred for 15 minutes and then acidified with a 1: 4 dilution of sulfuric acid to pH 2.5. The biomass and other insoluble ingredients are separated from the brown colored clear filtrate containing 3600 g / ml cephalosporin C by filtration.

000 ml of this filtrate was percolated through a 3 970 ml Ostion SP-1 resin cartridge at a mernor load of 1 h. The adsorbate residue was displaced from the cartridge by 2,000 ml of water and desorbed with unbound cephalosporin C with a 152% aqueous acetone solution.

3060 ml of an eluate containing 5,000 µg / ml of cephalosporins C are obtained, which is subjected to further purification.

The Ostion SP- used was regenerated with 12 L of a 252 N aqueous solution of acetone containing 22 NaOH. The cartridge was then washed with 2 L of 12-L sulfuric acid and finally with 12 L of water.

Example 2

Method as in Example 1, except that 5 mg of urea per ml of soil was added to the crude fermentation broth and cephalosporin C was eluted from the resin with 0.05 N aqueous NaOH. 5 450-ml of a clear filtrate containing 3 100 µg / ml of cephalosporin C is adsorbed onto 4000 ml of Ostion SP-1. This is desorbed from the sorbent with 0.05 N aqueous NaOH, yielding 82 2 of the 3 800 ml eluates. total amount of Cephalosporins C.

Example 3

Method as in Example 1, except that 30 mg of urea per ml of soil was added to the crude fermentation broth and the pH was adjusted to 2.9 (1: 4) with sulfuric acid. The clear filtrate obtained is subjected to further purification on Ostion SP-1.

Example 4

Method as in Example 1, except that 10 mg of urea per ml of soil and 0.5 per mille of anionic detergent Tergitol are added to the crude fermentation broth. The crude fermentation platform thus treated is filtered and the clear filtrate subjected to further purification.

Example 5

Method as in Example 1, except that 5 mg / ml urea and 5 per mille of anionic detergent Tergitol are added to the crude fermentation broth and the pH is adjusted to 4.0 with sulfuric acid (1: 4). The treated fermentation is then filtered and the filtrate subjected to further purification.

Claims (2)

Example 5 Method as in Example 1, except that 5 mg / ml of urea and 5 ml of anionic detergent Tergitol are added to the crude fermentation broth and the pH is adjusted to 4.0 with sulfuric acid / 1: 4 / . The treated suspension is filtered and the filtrate is further purified. SUMMARY OF THE INVENTION 1. A method for treating a fermentation process after biosynthesis of cephalosporin to the base is added, at pH 2.5-4.0 5-30 mg / ml urea, to give purification * 2. Process according to claim 1, characterized in that the anionic detergent is present in an amount of 0.5 - 5%. C, which is then filtered and subjected to the addition of Sevfrography. n. p- xivod 7. Most