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Three spore acid can promote three Alternaria the DON's Streptomyces p-carotene biosynthesis, while three Alternaria acid is a byproduct of the fermentation process can be extracted from the fermentation broth, is added to the fermentation broth back to increase the p-carrot hormone production, so the mechanism for the establishment of an economically viable extracted from the fermentation broth separation of the three spores acid process has great economic significance, the same time the the three spore acids promote the mechanism of action of p-carotene unclear, in recent years to study its role is increasingly attentions. HPLC assay of the acid content of the three spores, the chromatographic conditions were as follows: the column is a reversed phase C18 column (250 × 4.6 mm the 5μm, DiamodsilTM), the detection wavelength is 325nm, column temperature is 30 ° C, the mobile phase is methanol and water containing 0.1% TFA, using a gradient elution mode, pump A was 100% methanol, pump B is 0.1% TFA in ultrapure water, methanol promoted from the linearity of 50% 35min 70%, and the injection volume was 10μl. flow rate of 1.0ml/min. HPLC method than of UV spectrophotometry accuracy and precision, and quick and easy application of the detection process gradient elution not only shorten the analysis time, but also improves the separation. Liquid - liquid extraction method can be obtained the three spores acid purity of about 30%, NaHCO3 and chloroform extraction step is repeated with a slightly higher purity can be obtained three-spore acid, but not exceeding 40% purity. Sephadex LH-20 dextran gel chromatography three spore acid optimum conditions: eluent 40% aqueous methanol, flow rate 2ml/min, purity can be obtained the three spores acid is about 70% of . Silica gel column chromatography three Alternaria acid conditions: mobile phase chloroform: ethyl acetate: petroleum ether: acetic acid (V: V: V: V) = 14:3:5:0.5, when the sample volume is controlled at 100mg or less, the three spore acid purity of greater than 60%. Different macroporous resin three spore acid adsorption and desorption characteristics determine the sample solution SP850 best resin, to optimize the process of adsorption and desorption conditions: the the three spore acid concentration in the fermentation broth 15.30μg/ml pH 2, the adsorption velocity 6BV / h, the temperature was 25 ° C, and on the volume of the sample liquid is 30BV. The gradient elution program: first with deionized water, the addition of impurities, and then 3BV30% aqueous ethanol to remove most of the impurities, and finally eluting with 8BV60% aqueous ethanol most three spore acid elution rate for 9BV / h. After macroporous resin SP850 after three spore acid purity can reach 70%. For the first time three spore acid by preparative high-performance liquid chromatography separation conditions: 55% methanol aqueous solution as the mobile phase at a flow rate of 10ml/min, the sample volume should be controlled in less than 20mg and MS and 1H-NMR characterization of products structure confirmed three spore acid C. Studied in three spore acid and its structural analogues of p-ionone and abscisic acid is added of three spore DON's mold and \p-ionone and abscisic acid optimal added concentration were 9.10μM of 50μM and 0.909μM, p-carotene production increased by 70.4%, 47.2% and 43.4%. Studied the three spore Brad's mold \the gene carRA and carB expression of multiples is the most significant, respectively at 6h and 3h impact, increased 3.24 times and 9.93 times respectively; abscisic acid for carRA Genes and carB of the level of gene transcription in cultured 3h maximum , 1.97 times and 7.85 times respectively. p-ionone carRA Genes and carB of the level of gene transcription 6h and 3h maximum, 2.58 times and 6.10 times respectively.
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