CN1035333C - Production method of salinomycin and its sodium salt crystalline product - Google Patents

Abstract

The invention relates to a process for extracting salinomycin and its sodium salt from fermentation broth. Acidify the fermentation broth, add filter aid to filter, extract the filter residue with alcohol, filter, decolorize the filtrate with activated carbon, filter, take the filtrate to concentrate, and add water to obtain the product. The invention has the advantages of simple process, low cost, and low toxicity of the solvent used.

Description

Production method of salinomycin and its sodium salt crystalline product

The invention relates to a method for extracting antibiotic crystals from antibiotic biosynthesis products.

According to literature reports, the technological method of extracting salinomycin crystalline product from fermented liquid has:

(1) US Patent 3857948 (1974)

Filtrate the fermentation broth, extract the mycelia in the filter residue with organic solvents such as butyl acetate, acetone and chloroform after autolysis, concentrate the extract in vacuum and purify it with alumina column chromatography, and use ethyl acetate, n-butane Or the mixture of the two is eluted, the active part is collected, concentrated in vacuum and then chromatographed on Sephadex LH-20, the effective components are collected and concentrated to dryness to obtain salinomycin powder.

(2) Japanese Patent No. 53-148595:

Adjust the pH of the culture solution to 4.5-5 with diluted hydrochloric acid, stir at 60°C for 10 minutes, add 4% (W/V) filter aid, filter, and extract the filter residue twice with butyl acetate, and extract the solution with 5% carbonic acid Wash with 50 liters of sodium aqueous solution, evaporate to dryness under reduced pressure to obtain coarse powder, add 100 liters of n-hexane to dissolve the coarse powder, concentrate under reduced pressure to 40 liters, and refrigerate at 5°C to obtain crude salinomycin sodium. After chromatographic separation or repeated recrystallization with n-hexane, pure salinomycin sodium can be obtained.

(3) Antibiotics Reported in the fifth issue of 1982: add 4% diatomaceous earth to the fermentation broth, filter, extract the filter residue twice with 1/2 volume of butyl acetate, and fully wash the extract with 5% sodium bicarbonate solution , the aqueous phase was removed, and the organic layer was concentrated to an oil under reduced pressure. The oil was subjected to alumina column chromatography, eluting with ethyl acetate:methanol (100:0 → 100:2 → 100:20), collecting and combining the part of the eluate that had inhibitory activity against Bacillus subtilis, and concentrating under reduced pressure , the concentrate was chromatographed on a silica gel column, eluted with chloroform:methanol (100:0→100:2→100:20), and the components with Rf values similar to those of salinomycin were collected, concentrated to dryness, and reconstituted in aqueous acetone. Crystallization to obtain white salinomycin sodium salt crystals.

Most of the organic solvents used in the methods described above are toxic. Because the fermentation broth also contains a considerable amount of oil, other organic solvents except acetone also extract the oil into the organic phase when extracting salinomycin, and further oil removal is very troublesome in technology. Although this shortcoming can be avoided by adopting acetone, acetone has a low boiling point and has a large volatilization loss in the production process. In addition, the separation and repeated recrystallization by layer and column make the production process route too long, and it is difficult to realize industrial production.

The object of the present invention is to provide such a process for extracting salinomycin and its sodium salt from salinomycin fermentation broth, which has a simple and simple production process, low toxicity of solvents used, and high product purity and yield.

The present invention is realized in the following way: acidify the fermented liquid with acid, adjust pH 1-6, add 1-5% of various filter aids such as diatomite, oxalic acid, yellow and white salt, zinc sulfate, etc., filter under vacuum or pressure, discard The filtrate is removed, and the filter residue is extracted with a low-toxic organic solvent. The low-toxic organic solvent used is preferably alcohol or acetone, preferably alcohol, and then filtered under vacuum or pressure, the filtrate is taken, and a decolorizing agent such as activated carbon is added for decolorization. Filtrate under vacuum or pressure, take the filtrate to concentrate, and add appropriate amount of water to obtain salinomycin crystals. If the filtrate is concentrated, add alkali to adjust the pH to greater than 7, and then add water to obtain salinomycin sodium crystals.

Several examples are given below:

1. 100 liters of fermentation broth, the fermentation unit is 20000U/ml, add oxalic acid to adjust the pH=1, filter, add 2 times the amount of alcohol to the filter residue, stir for 2 hours, filter, add 1 kg of activated carbon to the filtrate, filter after stirring, filter and concentrate to about 10 liters, add 20 liters of water, stir and let it stand still, crystals precipitate out, filter to obtain white salinomycin crystals, dry weight 1400 grams, yield 70%.

2. Fermentation broth 100 liters, fermentation unit 2000U/ml, add hydrochloric acid to adjust pH = 1, add 4% diatomaceous earth, stir for 10 minutes, filter, add three times the amount of alcohol to the filter residue, stir for 2 hours, filter, add activated carbon 1 to the filtrate kg, filtered after stirring, concentrated the filtrate to about 10 liters, added 20 liters of water, stirred and left to crystallize, filtered to obtain white salinomycin crystals, dried weight 1450 grams, yield 72.5%.

3. Fermentation broth 100 liters, fermentation unit 25000U/ml, add phosphoric acid to adjust pH = 1, add yellow blood salt and zinc sulfate 0.1 kg each, heat to 60°C, stir for 30 minutes, filter, add three times the amount of alcohol to the filter residue and stir for 2 hour, filter, add 1 kg of activated carbon to the filtrate, filter after stirring, concentrate the filtrate to about 10 liters, adjust the pH of the concentrated solution to 10 with alkali, add 20 liters of water, leave to stand for crystallization after stirring, and filter to obtain white salinomycin sodium crystals. The drying weight is 1480 grams, and the yield is 74.1%.

The advantage of the invention is that the process is relatively simple, and qualified crystalline products can be obtained without chromatographic separation and repeated recrystallization. The industrial alcohol used is less toxic than the organic solvent used in the method introduced in the literature, and the price is more than twice as low, and the cost of the whole process is reduced by more than 50%.

Claims (6)

1. A process for extracting salinomycin and its sodium salt from salinomycin fermented liquid, characterized in that the fermented liquid is acidified with acid, the pH is adjusted at 1 to 6, filter aid is added, filtered, and the filter residue is used Extract with alcohol solvent or acetone, filter, take the filtrate and add decolorizing agent to decolorize, filter, concentrate the filtrate, add water to obtain salinomycin crystals, or add alkali to adjust the pH to greater than 7 after concentrating the filtrate, then add water to obtain salinomycin sodium crystals. 2. The process according to claim 1, the acid used is various organic acids or inorganic acids; it is advisable to adjust the acidity with pH1～6. 3. The process according to claim 1, characterized in that the filter aid used is diatomaceous earth, yellow blood salt or zinc sulfate. 4. The process according to claim 1, characterized in that the alcoholic solvent used is alcohol. 5. The process according to claim 1, characterized in that the decolorizer used is gac. 6. The process according to claim 1, characterized in that vacuum filtration or pressure filtration is used when filtering.