Tetracyclines (TC) are polyketide natural products that include a number of clinically important antibacterial drugs, such as oxytetracyclin (OTC) - a broad-spectrum antibiotic produced by the important industrial microorganism Streptomyces rimosus. Considering ever-growing need for new and economically affordable therapeutics, the pharmaceutical industry is continously optimizing the bioproceses and thus reducing the cost of antibiotic production. This also includes extraction and purification processes improvement. The aim of this work was to evaluate and test different processes of extraction of tetracyclines from the fermentation broth of Streptomyces rimosus culture. We used a genetically modified strain of S. rimosus, which produces two tetracycline intermediates during oxytetracycline biosynthesis - dehydro TC and dehydro OTC. S. rimosus, which produces OTC at high yield was used as a control. We carried out comparative anaysis where saturation of the fermentation broth using monosodium phosphate was compared to the methods where pH was corrected by HCl or NaOH, followed by the extraction with ethyl acetate in a ratio of 1: 1. Two types of amberlite resins were also added to the culture to achieve immediate adsorption of the produced tetracycline intermediated during the biosynthetic process. Tetracyclines were subsequently eluted from the resin with methanol. The presence of tetracyclines in both phases - aqueous and organic, was analyzed by thin layer chromatography, high pressure liquid chromatography and mass spectrometry coupled by liquid chromatography. The addition of XAD16N amberlite resins to the fermentation broth and the subsequent elution of tetracyclines from resins with methanol proved to be the most effective method. This method was also transferred to a larger volume and the methanol extracts were concentrated by evaporation under vacuum. The final extracts contained the target tetracycline intermediates in concentration of around 2 g/L dehydro TC and 0,4 g/L dehydro OTC.
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