Primary Study On The Solid-State Fermentational Condition And Mutation Breeding Of Strain Producing Antidiabetic Pigment

The strain F120, which can produce a large amount of Indolyl benzoquinone pigments(IBP), was isolated from forest soil of Chengdu. Oral administration of IBP resulted in significant glucose lowering in two mouse models of diabetes induced by Alloxan and Dexamethasone. IBP also exhibited an insulin-like effect on antidiabetic action. The result of acute toxicity test indicated that its oral LD50 value was more than 4g/kg on mice, so IBP was categorized as a low toxic substance. IBP is potential to become a new oral medicine in the treatment of diabetic patients. The IBP-producing ability of strain F120 was examined by rice solid-state medium, and its optimum cultivation conditions were acquired by orthogonal method. The optimal solid-state fermentational condition was that 500mL flask containing 30g wet local indica rice(24g dry local indica rice), pH6.4, 1% Tryptone(w/w) and 0.01% K+(w/w)being added in the culture medium, the flasks were incubated at 30℃ in incubator for 12 days. Under this condition, the yield of IBP reached 1.1315g/kg. In selection of IBP high-producing strain, the key step was to design the efficient method to select positive mutants. Based on the character that the biosynthesis ability of Indolyl benzoquinone pigments was in accordance with the ability of utilizing protein, the high efficient method was build for the primary selection of IBP high-producing strains, which core contents included the aspects: using casein peptone as the unique carbon resource and nitrogen resource , the primary selection plates were prepared. According to the ratio of diameter of transparent cycle and colony in casein peptone plates, we selected strains with large ratio. Using strain F120 as the starting strain, the mutant F120-Ⅰ16 was obtained after UV irradiation. The mutant F120-Ⅰ16 produced 0.479g/kg IBP and increased by 34.6% compared with the starting strain F120. Subsequently, using strain F120-Ⅰ16 as the starting strain, the mutant F120-Ⅱ31 was obtained after being mutagenized by NTG. The mutant F120-Ⅱ31 produced 0.621g/kg IBP and increased by 29.6% compared with the starting strain F120-Ⅰ16, and 74.4% compared with the strain F120.