| The soil pollution caused by uranium and other associated heavy metals is mainly slight and moderate in China.Phytoremediation is the preferred method for treating the slight but large area heavy metal p ollution.However,the method might be limited to application because of a lack of a safe,environmentally friendly and rapid degration technology for the hyperaccumulators.In order to obtain a scientific and reasonable heavy metal enrichment biomass disp osal method,the experiments were conducted to study the degradation progress of Ryegrass by screened bacteria and some combination of them under the treatment of uranium,lead and strontium.The key results are as follows:(1)In order to study the optimal conditions for determining the enzymatic activity of cellulase,the parameter of the determination of cellulase activity by DNS method is optimized by spectrophotometry.It is found that the absorbance value of reducing sugar coloring solution fluctuate at 480,490 nm,which indicate that they are not suitable as the determination wavelength.While the 5202 was close to 8)(62 on 520 nm,and the R2 is stably in about 0.9998 and the slope decreases as the wavelength increases in the range of 520 ~ 580 nm.The results show that the 520 nm is the suitable wavelength for testing the cellulase activity,at which the method could not achieve a good linear relationship,but also achieve better resolution.(2)For obtaining the dominant strain with good degradation characteristics,bacteria source samples are processed with restrictive rich cultivation,set training and separation purification,through which the strains that could directly or indirectly utilize Ryegrass can be got.And cellulose enzyme capacity of the strains could be qualitatively study by using transparent circle method.Next The ability to utilize ryegrass and heavy metal tolerance have been study by plate culture method.Eventually the dominant strain was identified by 16 S r DNA or 18 S r DNA.8 strains that can use ryegrass better and produce cellulose have been got through our research,which include 4 strains of bacteria(B3,B8,B9,B(KC)),2 strains of actinomycetes(A1,A3)and 2 strains of fungi(F1,F2).Investigation on heavy metal resistance shows that bacterial has the best resistance to heavy metals,actinomyce follow,fungi is the worst.After excluding a actinomycete with poor heavy metal resistanc,the identification results are as follows: A1? B3?B8?B9?B(KC)?F1?F2 is Streptomyces sp.?Brevibacillus brevis?Bacillus licheniformis?Bacillus badius?Bacillus subtilis subsp? Fusarium sp.?Microascus manginii respectively.The results showed that Bacillus subtilis was a kind of microorganism with strong resistance and good degradation performance.(3)To examine the degradation ability of the combined strains,the antagonism between strains,bacterial cellulase activity and degradation rate were investigated by coating method,DNS method and diff erence method respectively.The strains were combined by orthogonal experiment.The research finds only Brevibacillus brevis has minor inhibition to other bacteria.Microascus manginii has the best enzyme activity in fermentation early,and its CMC-Na enzyme activity is 13.97 u/m L.FPA enzyme activity is 9.76 u/m L.Microascus manginii has the bigest degradation of 9.63% in fungi and actinomycetes.Bacillus badius has the bigest degradation of 20.53% in bacillus.Through orthogonal m ethod to match the strains,the research finds that temperature levels is far greater than other factors.According to the different temperature of the orthogonal experiment again intuitive analysis,we concludes that fungal and actinomycete Streptomyces sp.,Fusarium sp.,Microascus mangi nii vaccination ratio is 2:2:3 and bacteria Bacillus licheniformis,Bacillus badius,Bacillus Subtilis subsp vaccination ratio is 1:1:1.The results showed that the enzyme activity was positively correlated with the degradation rate,the greater the enzyme activity of the strain,the greater the corresponding degradation rate.The different functionalities of the same microorganisms are different at different fermentation temperatures.Subsequent studies could be carried out by temperature step-by-step fermentation to maximize the effect Degradation of microbes.(4)In order to obtain highly degraded flora,the research uses the waste of straw,leaves,weeds and other soil as the bacteria source material to screen efficient flora,through restricted enrichment cultivation,heavy metal domestication and other technologies to filter samples and regarding paper decomposition capacity,ryegrass degradation rates and wood quality cellulose degradation rates as filter index.we obtained two flora that numbere as 1,6 respectively,which have a higher ryegrass degradation activity and a certain uranium and its associated heavy metal tolerance of bacteria.After 6d 38? solid fermentation,the degradation rates of flora 1 and flora 6 are 45.63% and 49.54% respectively,which of the degradation rates of lignocellulose are 66.21% and 76.30% respectively.Through the isolation and purification of the dominant strains in the two flora,the identification results of 16 S r DNA showed that the dominant strains are Bacillus subtilis.The results further show that Bacillus subtilis was a kind of microorganism with strong resistance and good d egradation performance.And the degradation efficiency of flora is significantly higher than that of single bacteria and combined bacteria,which indicated that the non-culturable microorganisms in the flora have a great effect on the degradation.(5)To improve the efficiency of ryegrass degradation,our research combined with the combination of bacteria and flora to obtain complex bacteria,and the degradation efficiency of ryegrass was investigated by differential method.The results show that the degradation rate of ryegrass under the condition of 38 ? is better than 28 ?.But the degradation rate of the lignocellulose under the condition of 28 ? was better than 38 ?.The degradation rate of the secondary inoculation was higher than that of the one inoculation.However,There was no significant difference in them.The results of research indicate that fungi and actinomycetes have higher degradation ability of wood fibers than bacteria,and the bacteria was inoculated with in the second stage of fermentation and bacteria quickly take advantage,so that the dissolution of lignocellulosic decomposition products has been improved.To sum up,the inoculation is carried out by adding highly effective combination agents to the highly effective flora in our research,and the fermentation was carried out by step-by-step fermentation accompanied by the change of temperature,which achieves the rapid degradation of uranium enrichment ryegrass.It can be seen that this study has obtained some key parameters of the rapid degradation technology for degrading the uranium-enriched ryegrass.Meanwhile,it has obtained the composite microbial system of which the biodegradable ability is better.It highly provides a theoretical and technical basis for the degradation of heavy metals-enrich biomass. |
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