Optimization Of Transformation Process Of An O-Dma-Producing Oxygen-Tolerant Mutant Bacterium And Making Foods Containing O-Dma

O-Desmethylangolensin(O-Dma)is a metabolite of daidzein with a variety of biological activities beneficial to humans,including the affinity of O-Dma to estrogen receptors and the antioxidant capacity is higher than that of its precursor substance daidzein.O-Dma also has many effects such as inhibiting cancer cell growth,maintaining bone density and lowering lipid.In the previous study,a strain named Aeroto-AUH-JLC108 was obtained,and it was able to convert the substrate daidzein to O-Dma in BHI medium in the presence of atmospheric oxygen.However,the high cost of BHI medium is not conducive to industrial production.Therefore,this study was designed to seek a medium with low cost and improved the bioconversion rate of O-Dma through medium optimization.First,experiments proved that the strain Aeroto-AUH-JLC108 could grow on LB medium and be capable of converting a small amount of daidzein to O-Dma,and the bioconversion rate is 5%,as a result,LB was identified as initial medium.Then,the components of carbon source,nitrogen sources,trace elements and phosphates in the initial medium were screened by a single factor test.The important components that affect the conversion rate were determined by the Plackett-Burman test,and experiments were designed to seek the addition of each important influencing at the highest bioconversion rate of O-Dma by the steepest climbing experiment.Finally,after the optimization of the response surface methodology,the optimal proportions of medium were sucrose 1.6%,yeast extract powder 5%,dipotassium phosphate 0.3%,sodium chloride 0.15%,magnesium sulfate 0.05%,and L-Cysteine 0.05%.The results indicated that the bioconversion rate reached 84% in optimized medium,which was higher than the bioconversion rate of 76% in BHI medium,while the cost was reduced by about 80%.Both culture medium and fermentation conditions play a key role in the process of converting the daidzein to O-Dma by the strain Aeroto-AUH-JLC108.Hence,after optimizing the culture medium,five important parameters were further optimized,including pH,temperature,inoculation,final substrate concentration,conversion time.Using the Plackett-Burman and single factor gradient test,three optimized parameters were preliminary determined.The pH was determined to be 7.5,the temperature was 39 ° C,and the final concentration of the substrate was 0.5 mM,wherein the conversion decreased with the increase of the final concentration of the substrate.Due to the toxicity of the substrate solvent methanol and DMF to the bacteria,the concentration of the fixed substrate in this study was 1.0 mM.Then,the optimum pH and temperature were further optimized by the response of the central composite design.Finally,when the concentration of the substrate was 1.0 mM,the pH value was 7.0,the temperature was 38.5 °C,while the inoculation amount was 10%,and the transformation time was 72 h,the bioconversion rate can reach 92%.Using the optimized medium and conditions,the fermentation system was expanded in a 5 L bio-fermenter,and the growth and transformation dynamics of the bacteria in the fermentation process were compared in the bio-fermenter and constant temperature incubator,respectively.The results showed that the growth and transformation state of the cells in the fermenter were better than those in the incubator.The maximum OD value of the zymotic fluid in the constant temperature incubator and bio-fermenter was 2.5 and 3.6,and the final conversion rate was 92% and 97%,respectively.The conversion rate is higher in the bio-fermenter than the incubator.It may be related to stirring and oxygen content.O-Dma has a variety of biological activities beneficial to the human body.This study also explored how to make O-Dma into snacks.According to the standard of compactness,lightness and simplicity,we chose milk and chocolate,and explored food ingredients and processing technology.Sensory evaluation was used as the criterion.When the ratio of skim milk powder and cotton white sugar in milk tablets was 2:1,the ratio of cocoa mass,cocoa butter and powdered sugar in chocolate was 4:2:4,the scores of sensory evaluation were the highest.The activity of O-Dma could be maintained after the highest temperature treatment of food preparation by HPLC detection.