| Neomycin sulfate is a salt of neomycin and sulfuric acid.Neomycin is a broad-spectrum antibiotic isolated from streptomyces Fortis 3535 and 3554 culture solution by Waksman ET AL in 1949.It has good curative effect on suppurative infection,otitis media,conjunctivitis,purulent rash,wound ulcer,etc.,it can also be used as an auxiliary medicine for other antibiotics,such as chloramphenicol,Erythromycin,etc..The latter can get better effect,so it is widely used in the field of medicine.In this paper,the stability and reliability of the analytical method of Neomycin were investigated based on the pre-column Derivation Method After six rounds of ARTP mutagenesis combined with streptomycin and ammonium sulfate resistance screening,a high-yield and ammonium-resistant strain was obtained,excellent spore-producing medium was selected and the fermentation was optimized by using organic nitrogen source reagent box combined with small assistant of organic nitrogen source analysis.Finally,the effect of Aeration on seed metabolism and the measurement of dissolved oxygen rate by dynamic method were studied in 50-liter seed pot,and the fermentation yield was increased by more than 30%through multi-scale study.The main contents and results of this paper are as follows:(1)stability and reliability of precolumn derivation method.The precision test,stability test and comparison with biological activity test were carried out for the determination of Standard Neomycin.The results showed that the RSD of Neomycin B peak area was 0.81%for the same sample six times in a row,which indicated that the method had high precision in determining Neomycin B titer,the neomycin b peak area of neomycin sulfate remained constant and stable in 0.5-9 H,and the titer of Neomycin B peak by pre-column derivatization was slightly lower than that by bioassay,which was 81%-85%of bioassay,and the data had a strong correspondence with the bioassay titer,and had a high coincidence with the bioassay titer.(2)breeding of ammonium-resistant and high-yielding strains.After six rounds of ARTP mutagenesis and resistance screening,five strains with good growth and large colony diameter were selected to prove their fermentation performance in 24-well plate.Finally,three strains(27-3#,29-6#,15-4#)were screened.The fermentative titer of the three strains was 20%higher than that of the original strain.The optimum concentration of ammonium sulfate in the fermentation medium was increased from 0.5-1.0%to 1.5-2.0%,and the genetic stability was good.(3)optimization of culture medium.Four kinds of spore-producing media were selected,and then the amount of calcium carbonate and ammonium sulfate added in the fermentation media was optimized.Sentence is too long,please supply a shorter sentence.(4)Effect of aeration rate on metabolism of seed pot.In Four 50L fermenters,the aeration ratio of the same batch of seeds was investigated at 35?,350 RPM and 35?,respectively,at 0.5 vvm,0.75 VVM,1.0 vvm,1.25 VVM,the changing trend of reducing sugar,total sugar,Ph,bacterial concentration,amino nitrogen,viscosity,Neomycin B yield and impurity C in 4 cans.The uniformity of the metabolism of the big and small seed pot under the same aeration ratio was investigated.The results showed that aeration ratio had little effect on the spore germination rate in the first 20 hours,higher aeration ratio(1.25 VVM)was beneficial to shorten the time of pH recovery,higher maximum bacterial concentration and lower impurity c after entering the logarithmic growth period.Conclusion:The ventilatory ratio in the first 20 hours was 0.5 VVM,and in the 21 hours the ventilatory ratio was 1.25 VVM.The best time for transplanting was 35-40 h,the metabolic rate of 31 seed pot was slower than that of 50L seed pot,and the maximum rising Ph was only 7.2,the time delay of reaching the maximum concentration was about 27h.The 50L scale fermentation of the optimized seed culture technology was verified,which was more than 50%higher than that before optimization. |
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