Preparation method of 2,6-difluorobenzamide by utilizing rhodococcus ruber
A technology of difluorobenzamide and Rhodococcus erythrococcus, applied in the biological field, can solve problems such as high requirements for production equipment and production conditions, low conversion rate and selectivity of the method, and harsh reaction conditions of the production process, so as to reduce production costs, The effect of high substrate tolerance and fast reaction speed
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[0028] Example 1:
[0029] A method for preparing 2,6-difluorobenzamide by using Rhodococcus rubrum, the preparation steps are:
[0030] ⑴Seed and fermentation culture: Inoculate the Rhodococcus ruber (CGMCC3090) strain into the seed culture medium, culture at 180r / min, 30℃, 28h to obtain the seed culture solution, and transfer it to the fresh fermentation at an inoculum of 3wt% Culture medium, 180r / min, 28°C, culture for 48h to obtain the bacterial fermentation broth:
[0031] Seed medium: glycerol 10g / L, peptone 5g / L, malt powder 3g / L, yeast powder 3g / L, NaCl1g / L, pH7.0;
[0032] Fermentation medium: glucose 20g / L, yeast powder 5g / L, MgSO 4 ·7H 2 O, KH 2 PO 4 , K 2 HPO 4 0.5g / L, urea 7g / L, NaCl 2g / L, CoCl 2 0.02g / L, sodium glutamate 1g / L, pH7.0. Among them, urea and inducer CoCl2 were sterilized separately for 3-5 minutes.
[0033] ⑵ Preparation of Rhodococcus rubrum suspension: centrifuge the above-mentioned bacterial cell fermentation broth, remove the supernatant, and use pH 7.5 ...
Example Embodiment
[0039] Example 2:
[0040] Steps (1), (2) and (4) in this embodiment are the same as those in Embodiment 1, and only step (3) is described below.
[0041] (3) Conversion of 2,6-difluorobenzonitrile: The concentration of substrate 2,6-difluorobenzonitrile is 1mol / L, the reaction volume is 10ml, the final concentration of Rhodococcus erythrococcus is 2.37g / L, and the final concentration is 1% methanol, the conversion system is 50mmol / LpH7.5KH 2 PO 4 -NaOH buffer solution, 30°C, 180r / min, 3h after transformation, prepare pure 2,6-difluorobenzamide.
[0042] After testing, the conversion rate of 2,6-difluorobenzonitrile in this step is 100%, and the selectivity to 2,6-difluorobenzamide is 100%.
Example Embodiment
[0043] Example 3:
[0044] Steps (1), (2) and (4) in this embodiment are the same as those in Embodiment 1, and only step (3) is described below.
[0045] ⑶ Conversion of 2,6-difluorobenzonitrile: the concentration of substrate 2,6-difluorobenzonitrile is 2.5mol / L, the reaction volume is 10ml, the final concentration of Rhodococcus rhodochrous cells is 2.37g / L, adding 1% Tween 80, the conversion system is 50mmol / LpH7.5KH 2 PO 4 -NaOH buffer solution, 30℃, 180r / min, after 24h conversion, prepare pure 2,6-difluorobenzamide.
[0046] After testing, the conversion rate of 2,6-difluorobenzonitrile in this step is 100%, and the selectivity to 2,6-difluorobenzamide is 100%.
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