Screening And Breeding Of Oxytetracycline Strain Tolerating High Dosage Of Antifoamer And Its Fermentation Technology

Oxytetracycline is manufactured by fermentation with streptomyces rimosus. Strains of streptomyces rimosus, play an important role in the fermentation process. In the Oxytetracycline fermentation process, the antifoam usually shows inhibition to the growth microorganism.Ultraviolet mutagenesis was employed and then strains that tolerate antifoam were screened.187 strains were obtained from the screening, and 83 higher fermentation efficiency strains in 10 times antifoam agent were obtained. Re-examination of the higher fermentation efficiency strains, and a strain that would tolerate high dosage of antifoam was gained eventually. No effect of the fermentation efficiency was shown when the strain that would tolerate high dosage of antifoam in 10 times antifoam concentration, but the fermentation efficiency of the control strain in 10 times antifoam concentration was decreased by 76.1%.The purity detection of the strain tolerating high dosage of antifoam was conducted, and 600 colonies of 99.5% purity were counted, with two bare colonies and a brown colony. This strain continuously was transplanted for four times, and the results showed that this strain showed relatively stable heredity. Metabolic stability of the strains was investigated, and the results show that the fermentation efficiency of the fermentation time, pH value, glucose content and soluble amino nitrogen content showed a better trend.The Plackett-Burman design, the steepest ascent experiment and the Box-Behnken central composite design were adopted in the response surface optimization of oxytetracycline in fermentation medium. Three main factors of the soybean meal, ammonium sulfate, and cobalt chloride were screened from eight fermentation medium components of the soluble starch, soybean powder, calcium carbonate, ammonium sulfate, sodium chloride, corn steep liquor, potassium dihydrogen phosphate, cobalt chloride. The best shake flask fermentation medium was:Soybean flour 2.75%, ammonium sulfate 0.347%, cobalt chloride 0.0131%, soluble starch 10%, calcium carbonate 0.5%, sodium chloride 0.5% potassium dihydrogen phosphate 0.05%, corn steep liquor 1%, and the antifoaming 0.01%. Three times of shaking flask fermentation parallel experiments was done according to the fermentation medium, the mean fermentation efficiency was 103.4% than before.The fermentation conditions of the oxytetracycline strain tolerating high dosage of antifoam were screened. And the best shake flask fermentation conditions were:seed age 20-24 h, initial pH 6.5, fermentation temperature 30℃, inoculum 10%, with volume of 40 mL, fermentation period 144 h. Three times of shaking flask fermentation parallel experiments was done according to the fermentation conditions, the mean fermentation efficiency was 112.4% than before.