Improvement Of Milbemectin Producing Strain By Rational Screening

Milbemectin (a mixture of milbemycin A₃ and A₄ and the ratio of A₃:A₄ was 30:70) was an outstanding acaricide. Milbemectin have fine application value and the vast marketplace prospect. In this paper, we attempt to improve their production through screening the high milbemectin-producing strains and optimizing of the fermentation conditions and put to use in production large scale having provided the reliable technological parameter.First, through purification, a stable high-yielding strain, H-103-8, was obtained, which was 4% higher than the original strain. It was regarded as an parent strain, which was treated with UV,NTG and EMS respectively, or firstly with UV and then with NTG. The results showed that the EMS was more sensitive to H-103-8. But NTG method was better, and NTG joint compound mutagenic effects were more obvious than those of a single mutagenic. Through the selection of the mutagenic, we reduced the efforts.In order to reduce the efforts, improvement of H-103-8 was performed by a rational screening strategy to increase its production. The parent strain named as H-103-8 was treated with NTG, and sodium acetate, sodium propionate and aminoacetic acid were used as screening agents successively. Finally a high producing strain H-107-34 was obtained and produced 854μg/mL milbemectin, which was 37% higher than the parent strain H-103-8. Then the strain H-107-34 was treated with NTG, EMS and compound mutagenic, and streptomycin, 2-deoxy-D-glucose, and 5-oxime-milbemectin were used as screening agents, successively. Finally, we obtained a high milbemectin production strain H—113-17 and its' production reached 1140μg/ml, which was 90% higher than that of the ancestral strain, H-101-4.Besides, the fermentation conditions of milbemectin were studied in shaking flask. First, we explored the influence of different carbon source, nitrogen source, inorganic salt, trace elements and precursor on milbemectin production. On single-factor experimental basis through statistical analysis.At the same time, we researched the temperature, pH value, fermentation time, the volume of vaccination, and found that the container of 25 mL, pH value of 7.2, the vaccination of 6%, 28℃, fermentation of 12 days, milbemectin titer was the highest. The productivity of the new process with H—113—17 reached 1280μg/mL, which was 112% higher than the original process with H-101-4. Furthermore, the tablism of milbemectin including living things amounts, PH value, general sugar contents and amino nitrogen contents was studies during the fermentation.