Breeding Of Natamycin-Producing Strain

The parameters of HPLC method and agar diffusion method to determinate natamycin bioassay were studied. HPLC method was decided to be the determination method of natamycin. Streptomyces gilvosporeus NO.59 was treated with DES and UV light. A mutant SG-56 with high natamycin yield was selected with streptomycin resistance screening. The medium and fermentation process of SG-56 were optimized.The parameters of HPLC were determined. The operating conditions were Phenomenex prodigy 5 μ ODS3 100 A(250mm× 4.60mm i.d., 5μm) at room temperature, methanol-water-phosphoric acid (85:15:0.15, V/V) as mobile phase with a flow rate of 1 .0mL/min and UV detection at 303nm. The results will not be affected by other components in the broth. The relative standard deviation was 0.24% (n=5), and the recovery was over 99%. The result was verified by comparing with agar diffusion method.NO.59 strain was screened by agar block method because of its excellent fermentation capability and was treated as original strain. The natamycin yield of NO.59 was 1.25g/L in shaking flask.At first, NO.59 was treated with DES. A streptomycin-resistant mutant S-71 was obtained by streptomycin resistance screening. The natamycin yield of S-71 was up to 1.64g/L. Secondly, S-71 was treated with UV light. A streptomycin-resistant mutant SG-56 was obtained by the same method. The yield of SG-56 was up to 2.41g/L.The optimum medium and fermentation conditions for natamycin production were demonstrated. The optimum medium consisted of(g/L):glucose 36, soya peptone 18 and yeast extract powder 4.5 and pH7.0. Inoculum volume was 2%. A 500mL flask containing 50mL medium was cultivated at 29℃ for 96h and the speed was 200r/min. The yield of natamycin under these conditions was 2.75g/L.