Study On Degradation Of The Phytoestrogen Isoflavone By Comamonas. Testosteroni

Environmental estrogen,having the similar effect with estrogen,can interfere with the synthesis of organisms hormone,secretion,transport, integration,active reaction,metabolism and digestion,and have negative impact on biological organisms for maintaining a normal dynamic, reproduction,growth and behavior.These chemicals,discharged to the environment mainly by the activities of human,undermine the stability and regulation of the body,and may affect a variety of sexual act, reproductive function,immune system,neural systems and so on,even if with a litter number.Phytoestrogens is a kind of plant chemical constituents having similar biological activity with animals estrogen,there are three categories mainly:isoflavones,lignans and coumestans.Plant estrogen show the activity of estrogen and anti-estrogen in vitro and in vivo,and it depends mainly on the phytoestrogens concentration in the cell culture medium.Presently,the content of isoflavone in local environment was increasing with the discharged wastewater and sewage of related enterprise;Isoflavones content in the feed additive is relatively high;and the human average consumption of isoflavones also is increasing.And the high isoflavone content in the environment and in vivo have a tremendous side effects to humanity and other organisms.However, the anti-estrogenic and estrogenic activity of plant estrogen have not been given adequate attention.3α-Hydroxysteroid dehydrogenase/carbonyl reductase(3α-HSD/CR)from Oomamonas testosteroni is a key enzyme involved in the degradation of steroids and xenobiotic.Comamonas testosteroni are able to use steroids (testosterone,progesterone,bile acid)and other substances as sole carbon and energy sources.Complete assimilation of steroids is achieved through a complex pathway involving many enzymatic steps of oxidation responsible for the breakdown of the steroid nucleus.Comamonas testosteroni therefore play a significant role in the bioremediation of these stablely hormonally active compounds in the environment.Firstly we determine the methods of extraction and detection of soybean isoflavones,according to experiments of temperature,time,inoculum concentration,static and oscillating,we study the best conditions of degradation soybean power and soybean products by several strains.The main results are as follows:1.With different organic solvents as extracting solutions,ultrasonic extraction of soybean isoflavones at different temperatures,detect isoflavone concentration with HPLC.The results showed that ultrasonic extraction twice with 70%methanol at 70℃,25 min in frist time;20 min in second time,chromatographic conditions,column:C₁₈column; Detector:Waters 2996 PDA;mobile phase:methanol(A),0.25%acetic acid water(B),0 min A increase from 25%gradient,increasing curve 6;detection time:30 min;temperature:30℃;velocity:1 mL/min,soy isoflavones can be isolated well.2.In the processing of temperature and time influenceing on the degradation of isoflavones in soybean powder,the degradation rate of C.t,Ca3,Cp4,Cp7,P8 and PBBP increase with the temperature raise form 4℃,15℃,25℃to 30℃respectively,the degradation rate of strains are maximum at 30℃.The degradation rate increase from the the 5th day,the 10th day,the 15th day to the 20th day,at the 20th day reachs maximum.The degradation rate of Ca3 was 93.9%at the 20th day,30℃.Variance analysis show that:four levels of the temperature and the time reach a significant difference.3.In the experiment of degradation rate for different soybean varieties, the results show that:the degradation rate of Weilaimushi is the largest,followed by the Dongnong 410.As the degradation weight,the higher of the isoflavone content is,the higher rate of degradation rate.Variance analysis show that the time and strains is significant difference:strains and varieties is very significant difference.4.The degradation rate is very much related to bacterial concentration,it show that the degradation rate of vaccination of 800μL and 500μL,500μL 200μL and the reach a very significant difference,but the degradation rate of vaccination of 1000μL and 800μL is not significant differences,so vaccination of 800μL is the most suitable.5.With the two saltmediums,the MS₂ has better effect than MS₁.When the concentration of isoflavone increase from 0.05 mg/mL,0.15 mg/mh,0.65 mg/mL to 0.90 mg/mL,the degradation rate of bacterias also rise,but between 0.65 mg/mL and 0.90 mg/mL show no significant differences, among different strains,Ca3 demonstrate the best ability of degradation.6.C.t and Ca3 can grow from the range pH 5 to pH 9,and have the best degradation ability in the neutral condition of pH 7;the degradation rate between pH 6and pH 8,pH 5 and pH 9 was not significant different.7.The degradation of soybean products,the degradation rate of 30℃is higher than 40℃,15℃and 4℃,and reachs very significant differences,it showes the higher temperature is not good to enhance the degradation rate;compared with the stillness,oscillation can increase the degradation rate of strains.