CN104762288A - Bagasse immobilized combination bacterium and application thereof in soil polyaromatic hydrocarbon phenanthrene pollution restoration - Google Patents
Bagasse immobilized combination bacterium and application thereof in soil polyaromatic hydrocarbon phenanthrene pollution restoration Download PDFInfo
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Abstract
The invention discloses a bagasse immobilized combination bacterium and an application thereof in soil polyaromatic hydrocarbon phenanthrene pollution restoration. The phenanthrene is taken as a model of the polyaromatic hydrocarbon bagasse immobilized combination bacterium for restoring the soil polyaromatic hydrocarbon, the biologic material bagasse is taken as a carrier for immobilizing the trichoderma harzianum fungi strain ZG1 and the Klebsiella bacterium ZG2, and the bagasse immobilized fungus ZG1 and the bacterium ZG2 combination bacteria are utilized for restoring the soil polluted by polyaromatic hydrocarbon phenanthrene. The carrier biologic material bagasse is low in cost, the process of restoring soil polluted by polyaromatic hydrocarbon phenanthrene by using the bagasse immobilized combination bacteria ZG1+ZG2 is simple and environmentally friendly without causing secondary pollution, simultaneously the bio-availability of the polyaromatic hydrocarbon is increased and the efficiency of the micro-biological degradation is increased.
Description
[technical field]
The present invention relates to a kind of bagasse immobilization combination bacterium and the application in soil Phenanthrene pollution amelioration thereof, belong to environment pollutant biological treatment technical field.
[background technology]
Polycyclic aromatic hydrocarbons (PAHs) is the persistence organic pollutant that a class contains more than 2 phenyl ring.The pollution of Polycyclic Aromatic Hydrocarbonat Existing in Environment is mainly derived from the burning of the fuel such as coal, oil, Sweet natural gas and the smelting of oil and traffic pollution etc.China is the discharge big country of PAHs, and the amount of 16 kinds of PAHs pollutents of EPA's priority acccess control of China in 1980 discharge is 18000 tons, and by 2003, oneself was more than 25300 tons, and 2004 up to 114000 tons, account for 20% of world's total amount.P in soil AHs pollutes and mainly uses from atmosphere dried wet deposition, sewage irrigation, fertilizer and pesticide and straw burning etc.And sewage irrigation is China P in soil AH
sthe main mode polluted.Because PAHs has half volatile, low-solubility and difficult for biological degradation, be often adsorbed on soil particle, long-term in soil exist and accumulate in a large number, and therefore, soil carries the PAHs of in environment 90%.Current China's economic is in high-speed development period, also can increase further the demand of coal and oil, it can thus be appreciated that in soil, PAHs pollution also can increase the weight of further within the certain period from now on.
The a lot of Soils In The Region of China has polycyclic aromatic hydrocarbons to detect, and some areas soil is polycyclic aromatic hydrocarbons contaminated quite serious.2012, Cao Yunzhe etc. are based on bibliographical information result, the content distribution feature of 16 kinds of PAHs and Σ PAHs in the topsoil of 14 provinces of China, 3 municipalities directly under the Central Government, 2 autonomous regions and the Hong Kong Special Administrative Region is analyzed, the mean value of result soil Σ PAHs is 3654.97 μ g/Kg, and content highest zone reaches 151.6mg/Kg.
PAHs has chronic toxicity and carcinogenic, teratogenesis, mutagenic " three cause " effect, and the PAHs of the medium-term and long-term a large amount of accumulation of soil, by the transmission of food chain, forms great threat to human health.Therefore, the polycyclic aromatic hydrocarbons contaminated harm caused ecotope and HUMAN HEALTH of removing soil has become one of problem in the urgent need to address at present.
The seriousness polluted in view of soil polycyclic aromatic hydrocarbons (PAHs) and hazardness, the polycyclic aromatic hydrocarbons contaminated recovery technique of soil also receives extensive concern in recent years.Microorganism remediation technology have safety, effectively, the feature of cheap and non-secondary pollution, therefore, adopt microbiological deterioration PAHs to be considered to remove the most effectively one of section of having of PAHs pollutent in environment.In recent years, report the microorganism of many efficient degradation PAHs both at home and abroad, but these efficient degrading bacterias exist the environment that single bacterial strain is less competitive than indigenous microorganism and disposal field is in actual applications unsuitable for the problems such as the growth of efficient degrading bacteria, therefore, needed to be degraded by the mode of fungi-bacterium combined degraded the PAHs of difficult degradation.
Immobilized microorganism technique mainly refers to the area of space limited by Microorganism incubation free for dispersion by the method for physics or chemistry, to improve the concentration of microorganism cells, makes it keep higher biological activity and the method recycled.Microbe density is large, speed of response is fast and resistance to environmental impact feature owing to having for immobilized microorganism, therefore, this technology is expected to overcome the harmful competition that is added to free efficient degradation microorganism in reparation scene and indigenous bacterium or is difficult to the problem that conforms, realizes the effective repairing polluted soil of efficient degrading bacteria.Immobilized microorganism renovation of organic pollution soil is just receiving increasing concern for this reason.
Be used as microbial immobilized carrier at present and mostly be the higher organic carrier of cost, as glutaraldehyde, alginate calcium, sodium alginate, polyvinyl alcohol and polyacrylamide gel etc., be difficult to the requirement adapting to suitability for industrialized production.
In addition, repair in PAHs contaminated soil immobilized microorganism, associating strain degradation ability is higher than single bacterial classification, and employing bagasse immobilization fungi-bacterium composite bacteria rehabilitating soil is polycyclic aromatic hydrocarbons contaminated huge application potential.
And luxuriant and rich with fragrance (phenanthrene) is modal polyaromatic hydrocarbon pollutant in environment, it is also the mark of tobacco tar and motor vehicle exhaust polycyclic aromatic hydrocarbons.General higher at the concentrations of contaminated soil China and Philippines.
[summary of the invention]
The object of the invention is to overcome the difficult problem that the fixation support cost of existing immobilized microorganism degrading polycyclic aromatic hydrocarbons phenanthrene existence is higher, the bagasse immobilization combination bacterium that the degraded soil Phenanthrene providing a kind of energy efficient, safe and cheap pollutes.
Another object of the present invention is to provide the application of above-mentioned bagasse immobilization combination bacterium in soil Phenanthrene pollution amelioration.
The present invention to achieve these goals, by the following technical solutions:
A kind of bagasse immobilization combination bacterium, it is characterized in that employing bagasse is the combination of carrier immobilized fungus and bacterium, wherein said fungi is trichoderma harziarum Pseudomonas, and described bacterium is Klebsiella.
Bagasse is biological material, the bagasse carrier of biological material bagasse immobilized microorganism system not only can shield the competition of indigenous microorganism and the infringement of unfavorable edaphic condition and then ensure the micro-raw growth of its load, polycyclic aromatic hydrocarbon pollutant also in adsorbable Enriching soil, thus increase the bioavailability of polycyclic aromatic hydrocarbons, improve the efficiency of fixing microbiological deterioration polycyclic aromatic hydrocarbons in its surface.Biological material bagasse is agricultural wastes, containing nutritive elements such as abundant C, N; Natural organic matter can be discharged to soil, improve soil physico-chemical property, promote soil microbial activities and quantity, strengthen PAHs and remove; Very strong affinity is had to microorganism; Can natural degradation in soil, with after the simple environmental protection of process, can not secondary pollution be caused; And it is cheap.
Therefore, the technology that biological material bagasse immobilization fungi-bacterial remediation soil is polycyclic aromatic hydrocarbons contaminated, to elimination Pollution of Polycyclic Aromatic Hydrocarbons in Soil, avoids the polycyclic aromatic hydrocarbons contaminated transmission by food chain to have very important meaning to the threat that human health is formed.
Trichoderma harziarum Pseudomonas (Trichoderma harzianum) fungal bacterial strain (hereinafter referred to as ZG1) in the present invention and Klebsiella (Klebsiella sp) bacterium (hereinafter referred to as ZG2) are cultivated through selective enrichment by the soil near petroleum chemical plant, separation and purification obtains.ZG1+ZG2 can be the luxuriant and rich with fragrance degradation bacteria that sole carbon source carries out growing with phenanthrene.
Fungus and bacterium in the present invention is prepared into fungi bacteria suspension and Bacteria suspension respectively through cultivation in advance when being inoculated in bagasse carrier, cultivates preparation and obtain especially by following steps:
Fungal bacterial strain ZG1 after purifying and bacterial isolates ZG2 is inoculated in 50mL liquid nutrient medium respectively, and pH is adjusted to 7.0, in 30 DEG C, on the shaking table of 150r/min, shaking culture is maximum to the biomass of microorganism, generally needs 48h, get the bacterium liquid of 4mL respectively in centrifuge tube, centrifugal 10min under 4000r/min, then pours out supernatant liquor in centrifuge tube, and supplementary phosphoric acid buffer continues centrifugal 10min, repetition like this 3 times, making bacteria suspension, to be placed in 4 DEG C of refrigerators for subsequent use.
In the present invention, in 30 DEG C on bagasse carrier fungi bacteria suspension and Bacteria suspension being inoculated in respectively the process of inorganic salt liquid nutrient solution, in the shaking table of rotating speed 150rpm, shaking culture is to biomass maximum and obtained bagasse immobilization combination bacterium ZG1+ZG2.
Fungi bacteria suspension in the present invention and Bacteria suspension are 1:1:2 ~ 6 (ml/ml/g) with the amount ratio of the bagasse processed, preferred fungi bacteria suspension and Bacteria suspension are 1:1:3 (ml/ml/g) with the amount ratio of the bagasse processed, and make luxuriant and rich with fragrance degradation efficiency the highest.
In the present invention with fungi bacteria suspension and Bacteria suspension obtain bagasse immobilization combination bacterium step be:
The bagasse carrier taken, add after clean oven dry shreds in Erlenmeyer flask, aseptic inorganic salt liquid nutrient solution is added after sterilising treatment, add fungi bacteria suspension and Bacteria suspension more respectively, in 30 DEG C after mixing sealing, shaking speed is that in 150rpm shaking table, in shaking culture to bagasse carrier, microbial biomass reaches maximum and namely obtains bagasse immobilization combination bacterium ZG1+ZG2, and wherein said inorganic salt liquid nutrient solution and the amount ratio of bagasse carrier are 10:3 (mL/g).
The application of a kind of above-mentioned bagasse immobilization combination bacterium ZG1+ZG2 in soil Phenanthrene pollution amelioration.
1, be the soil of 84.35mg/kg for luxuriant and rich with fragrance starting point concentration, after bagasse immobilization combination bacterium ZG1+ZG2 and free combination bacterium ZG1+ZG2 process 25d, luxuriant and rich with fragrance degradation rate is respectively 52.89% and 94.02%, shows that bagasse immobilization combination bacterium G1+ZG2 is better than free bacterium greatly to the degradation effect of phenanthrene.
2, after bagasse immobilization combination bacterium ZG1+ZG2 process 25d, the degradation rate of non-sterilized soil China and Philippines reaches 94.99%, and the degradation rate of sterilized soil China and Philippines is 90.81%, no matter soil whether sterilizing, the degradation rate of soil China and Philippines all can reach more than 90%, therefore, the impact of soil microorganisms on bagasse immobilization combination bacterium degraded soil China and Philippines is less.
When 3, preparing bagasse immobilization combination bacterium, best bacteria suspension add-on is that every 15g soil adds 1mL combination bacterium ZG1+ZG2 (wherein each 0.5mL of ZG1 and ZG2).The bagasse immobilization combination bacterium process starting point concentration adopting best bacteria suspension add-on to prepare is the phenanthrene-polluted soil of 84.65mg/kg, and luxuriant and rich with fragrance degradation rate reaches 94.55%.
4, the heavy metal ion Cd in soil
2+and Pb
2+to bagasse immobilization combination bacterium ZG1+ZG2 process soil, all there are certain influence in China and Philippines, even if but there is heavy metal ion Cd in soil
2+and Pb
2+, bagasse immobilization combination bacterium still can reach more than 80% to the degradation rate of soil China and Philippines.These illustrate that bagasse immobilization not only has the ability of stronger degraded soil China and Philippines, and have good adaptive capacity to environment.
Preferably, bagasse immobilization combination bacterium ZG1+ZG2 consumption is with the contaminated soil in its 10 times of weight of bagasse carrier 1 weight part process.
Enter preferably further, Phenanthrene contaminated soil first carries out the process of duplicate removal metal ion before repairing with bagasse immobilization combination bacterium, especially Cd
2+and Pb
2+transformatin.
The present invention, relative to prior art, has following advantage:
Bagasse immobilization combination bacterium used carrier biological material bagasse cost of the present invention is low, simple environmental protection when bagasse immobilization combination bacterium ZG1+ZG2 rehabilitating soil Phenanthrene pollutes, secondary pollution can not be caused, the bioavailability of polycyclic aromatic hydrocarbons can be improved simultaneously, improve the efficiency of microbiological deterioration polycyclic aromatic hydrocarbons.
[accompanying drawing explanation]
Fig. 1 is the effect of free combination bacterium and bagasse immobilization combination bacterium degraded soil China and Philippines;
Fig. 2 is that sterilizing and non-sterile soil combine the impact of bacterium degraded soil China and Philippines to bagasse immobilization;
Fig. 3 is that difference adds the impact of bacterium amount on bagasse immobilization combination bacterium degraded soil China and Philippines;
Fig. 4 attaches most importance to Metal Ions Cd
2+and Pb
2on the impact of bagasse immobilization combination bacterium degraded soil China and Philippines;
Wherein, the transverse axis in Fig. 1-Fig. 4 is all time (d) axle, and the longitudinal axis is all degradation rate (%) axle;
In Fig. 1, the meaning of each bar curve representative is:
--control treatment,
The degradation rate of-zero-free combination bacterium process soil China and Philippines,
The degradation rate of-△-immobilization combination bacterium process soil China and Philippines.
In Fig. 2, the meaning of each bar curve representative is:
-■-non-sterile soil control treatment,
-● the degradation rate of-non-sterile soil China and Philippines,
-△-sterile soil control treatment,
The degradation rate of — ╳-sterile soil China and Philippines.
In Fig. 3, the meaning of each bar curve representative is:
— ╳-process 1 (control treatment),
-● the degradation rate of soil China and Philippines in-process 2,
The degradation rate of soil China and Philippines in--process 3,
The degradation rate of soil China and Philippines in-▼-process 4.
In Fig. 4, the meaning of each bar curve representative is:
-■-control treatment,
-●-add the degradation rate of bacterium process soil China and Philippines,
-▲-Cd
2+the degradation rate of bacterium process soil China and Philippines is added when existing,
-▼-Pb
2+the degradation rate of bacterium process soil China and Philippines is added when existing.
[embodiment]
A kind of bagasse immobilization combination of the present invention bacterium ZG1+ZG2, being that carrier immobilized trichoderma harziarum Pseudomonas fungi ZG1+ Klebsiella bacteria ZG2 combines bacterium by adopting biological material bagasse, obtaining particular by following technical scheme:
The soil gathered near petroleum chemical plant carries out enrichment culture, separation and purification goes out ZG1 and ZG2, carry out domestication respectively cultivate and be prepared into ZG1 bacteria suspension and ZG2 bacteria suspension, then biomass ZG1 bacteria suspension and ZG2 bacterial suspension inoculation being cultured on the bagasse carrier of inorganic salt liquid nutrient solution process microorganism is maximum;
Wherein cultivate preparation ZG1 bacteria suspension and the condition of ZG2 bacteria suspension is: pH is adjusted to 7.0, in 30 DEG C, on the shaking table of 150r/min after shaking culture 48h, isolating bacteria suspension with the centrifugation of 4000r/min, to be placed in 4 DEG C of refrigerators for subsequent use;
The condition wherein preparing bagasse immobilization combination bacterium is: inoculate ZG1 bacteria suspension and ZG2 bacteria suspension after bagasse carrier by fungi bacteria suspension and Bacteria suspension and amount ratio 1:1:2 ~ 6 (ml/ml/g) of the bagasse processed, in 30 DEG C, shaking speed is shaking culture 2d in 150rpm shaking table, biomass to microorganism reaches maximum, and presentation is that bagasse carrier covers with bacterium.
The present invention's various substratum used is as follows:
Liquid nutrient medium: extractum carnis 5g, peptone 10g, glucose 5g, NaCl 5g, distilled water l000mL, pH 7.5.Solid medium need add 20-25g agar.
Minimal medium (inorganic salt solution): K
2hPO
40.5g, KH
2pO
40.5g, NaCl 0.2g, MgSO
40.2g, NH
4nO
31.0g, trace element solution 10mL, distilled water l000mL, pH 7.2-7.5.
Trace element solution: FeSO
40.1g/L, MnSO
40.1g/L, ZnSO
40.1g/L, Na
2moO
40.01g/L, CaCl
20.1g/L, MgSO
43g/L, CuSO
40.1g/L.
Below in conjunction with specific embodiment, the present invention is described in further detail:
Embodiment 1: the effectiveness comparison of free combination bacterium ZG1+ZG2 and bagasse immobilization combination bacterium ZG1+ZG2 degraded soil China and Philippines
Bacterial strain ZG1 and ZG2 after purified is inoculated in 50mL liquid nutrient medium respectively, and pH is adjusted to 7.0, in 30 DEG C, shaking culture 48h on the shaking table of 150r/min, the biomass to microorganism reaches maximum, get the bacterium liquid of 4mL respectively in centrifuge tube, centrifugal 10min under 4000r/min, then pours out supernatant liquor in centrifuge tube, mends appropriate phosphoric acid buffer and continues centrifugal 10min, repetition like this 3 times, making bacteria suspension, to be placed in 4 DEG C of refrigerators for subsequent use.
Take from Agricultural University Of South China's arboretum for examination red earth, its quality is silt loam, and pH value (water/soil=2.5/1) is 5.35, and organic matter is 7.89g/kg, nitrogen, phosphorus, potassium be other 0.71,0.72,2.61g/kg, phenanthrene does not detect.
Cross at 500g in the soil of 2mm sieve and add the luxuriant and rich with fragrance acetone soln that the luxuriant and rich with fragrance content of 50mL is 2000mg/L, stir, cross the mixing with soil that 2mm sieve after room-dry again with 500g even, namely obtain manual simulation's phenanthrene-polluted soil.Camera bellows cultivates contaminated soil 4 weeks, and the starting point concentration measuring contaminated soil China and Philippines before experiment is 84.35mg/kg.
Bagasse immobilization combination bacterium ZG1+ZG2 preparation: take the optimum carrier bagasse 1.5g selected respectively, clean oven dry shreds and adds in 250mL Erlenmeyer flask respectively afterwards, prepare empty 250mL Erlenmeyer flask to be used as the cultivation of free microorganism simultaneously, be placed in the high-pressure sterilizing pot sterilizing 25min of 1.5Mpa together, be transferred to rapidly on aseptic operating platform after pan gas drops subject to sterilization, sterilizing cool to room temperature under ultraviolet lamp, then in each Erlenmeyer flask, the aseptic inorganic salt liquid nutrient solution of 5mL is respectively added, 0.5mL ZG1 and 0.5mL ZG2 bacteria suspension is added respectively in each Erlenmeyer flask again with liquid-transfering gun, mixing, sealing bottleneck, labelled, be placed in 30 DEG C, shaking speed is shaking culture in 150rpm shaking table, add 15g phenanthrene-polluted soil after 2d and carry out reparative experiment.
Test arranges following 3 process: process 1: control treatment (not adding bacterium); Process 2: free combination bacterium ZG1+ZG2 process (adding each 0.5mL of ZG1 and ZG2 bacteria suspension); Process 3: immobilization combination bacterium ZG1+ZG2 process (bagasse 1.5g+0.5mL ZG1 and 0.5mL ZG2 bacteria suspension).
Have 15 beakers under often kind of process, respectively at the 5th, 10,15,20, the residual quantity of 25d sampling and measuring phenanthrene, often kind of process 3 is parallel.The water ratio of tested soil in each process is kept to be about 15% in test.After process terminates, measure the residual quantity of each process contaminated soil China and Philippines and calculate luxuriant and rich with fragrance degradation rate, for the soil of equal luxuriant and rich with fragrance pollution level, immobilized bacterium and free bacterium repairing effect are as shown in Figure 1.
Degradation rate luxuriant and rich with fragrance is as seen from Figure 1 respectively in control treatment, the process of free combination bacterium, the process of immobilization combination bacterium: 9.65%, 52.89%, 94.02%, the process of immobilization combination bacterium makes luxuriant and rich with fragrance degradation rate specific ionization combination bacterium process exceed nearly 42 percentage points.This illustrates that immobilization combination bacterium is better than free combination bacterium greatly to the degradation effect of phenanthrene.
Embodiment 2: sterilizing and non-sterile soil are on the impact of bagasse immobilization combination bacterium ZG1+ZG2 degraded soil China and Philippines
Bacteria suspension is with embodiment 1, and the added bacterium of each process is bagasse immobilization combination bacterium ZG1+ZG2, and prepare with embodiment 1 for examination soil and bagasse immobilization ZG1+ZG2, the starting point concentration measuring contaminated soil China and Philippines before experiment is 82.05mg/kg.
The present embodiment arranges following 4 process:
Process 1: non-sterilized soil adds bacterium process (adding bagasse immobilization combination bacterium ZG1+ZG2)
Process 2: non-sterilized soil control treatment (not adding bacterium)
Process 3: sterilized soil adds bacterium process (adding bagasse immobilization combination bacterium ZG1+ZG2)
Process 4: sterilized soil control treatment (not adding bacterium)
Have 15 beakers under often kind of process, respectively at the 5th, 10,15,20, the residual quantity of 25d sampling and measuring phenanthrene, often kind of process 3 is parallel.The water ratio of tested soil in each process is kept to be about 15% in test.
After process terminates, measure the residual quantity of each process contaminated soil China and Philippines and calculate luxuriant and rich with fragrance degradation rate, for the soil of equal luxuriant and rich with fragrance pollution level, the degradation rate of sterilizing and non-sterile soil China and Philippines as shown in Figure 2, no matter whether soil disinfection adds 10d before bacterium process, luxuriant and rich with fragrance degradation rate slowly increases, and degradation rate luxuriant and rich with fragrance after 10d increases rapidly, and after 20d, degradation rate change tends towards stability.After adding bagasse immobilization combination bacterium ZG1+ZG2 process 25d, the degradation rate of non-sterilized soil China and Philippines reaches 94.99%, higher than the degradation rate 90.81% of sterilized soil China and Philippines, no matter soil whether sterilizing, the degradation rate of soil China and Philippines all can reach more than 90%, therefore, sterilizing and non-sterilizing affect less on the degradation rate of soil China and Philippines.
Embodiment 3: different bacteria suspension add-on is on the impact of bagasse immobilization combination bacterium ZG1+ZG2 degraded soil China and Philippines
Bacteria suspension is with embodiment 1, and the added bacterium of each process is bagasse immobilization combination bacterium ZG1+ZG2, and for examination soil with embodiment 1, the starting point concentration measuring contaminated soil China and Philippines before experiment is 84.65mg/kg.
Bagasse immobilization ZG1+ZG2 combines the preparation of bacterium: the optimum carrier bagasse 1.5g selected taking equal in quality respectively, clean oven dry shreds and adds in 250mL Erlenmeyer flask respectively afterwards, prepare the 250mL Erlenmeyer flask of the sky of same number to be used as the cultivation of free microorganism simultaneously, be placed in the high-pressure sterilizing pot sterilizing 25min of 1.5Mpa together, be transferred to rapidly on aseptic operating platform after pan gas drops subject to sterilization, sterilizing cool to room temperature under ultraviolet lamp, the inorganic salt liquid nutrient solution that 5mL is aseptic is respectively added in each Erlenmeyer flask, then different amount bacteria suspension is got in each Erlenmeyer flask by 4 different treatment liquid-transfering guns, mixing, sealing bottleneck, labelled, be placed in 30 DEG C, shaking speed is shaking culture in 150rpm shaking table, add 15g phenanthrene-polluted soil after 2d and carry out reparative experiment.
The present embodiment arranges 4 process:
Process 1: control treatment (when preparing bagasse immobilization combination bacterium, bacteria suspension add-on is 0);
Process 2: when preparing bagasse immobilization combination bacterium, bacteria suspension add-on is 0.25mL ZG1 and 0.25mL ZG2;
Process 3: when preparing bagasse immobilization combination bacterium, bacteria suspension add-on is 0.5mL ZG1 and 0.5mL ZG2;
Process 4: when preparing bagasse immobilization combination bacterium, bacteria suspension add-on is 0.75mL ZG1 and 0.75mL ZG2.
Have 15 beakers under often kind of process, respectively at the 5th, 10,15,20, the residual quantity of 25d sampling and measuring phenanthrene, often kind of process 3 is parallel.The water ratio of tested soil in each process is kept to be about 15% in test.After process terminates, measure the residual quantity of each process contaminated soil China and Philippines and calculate luxuriant and rich with fragrance degradation rate, for the soil of equal luxuriant and rich with fragrance pollution level, difference adds the degradation rate of soil China and Philippines after the process of bacterium amount as shown in Figure 3.
As seen from Figure 3, after adding bacterium process 10d, the degradation rate of soil China and Philippines all increases sharply, and degradation rate change luxuriant and rich with fragrance after 20d eases up.Along with the increase of bacteria suspension add-on, the degradation rate also corresponding increase of soil China and Philippines, but when bacteria suspension add-on continue to be increased to total amount be 1.5mL time, although luxuriant and rich with fragrance degradation rate still increases to some extent, but very limited, with to add bacterium total amount 1mL phase difference little, this is due to the increase along with bacteria suspension add-on, carbon source relative deficiency needed for microorganism growth, causes effective bacterium source similar.Therefore in actual application, for cost-saving, the bacteria suspension add-on of process 3 is optimal addn.
Embodiment 4: heavy metal in soil ion Cd
2+and Pb
2on the impact of bagasse immobilization combination bacterium ZG1+ZG2 degraded soil China and Philippines
Bacteria suspension is with embodiment 1, and the added bacterium of each process is bagasse immobilization combination bacterium ZG1+ZG2, for examination soil and bagasse immobilization combination bacterium ZG1+ZG2 preparation with embodiment 1, but the PbC1 respectively containing 0.01% in sterilizing inorganic salt liquid substratum wherein
2and CdC1
2, the starting point concentration measuring contaminated soil China and Philippines before experiment is 85.25mg/kg.
The present embodiment arranges following 4 process:
Process 1: control treatment (not adding bacterium)
Process 2: add bacterium process (adding bagasse immobilization combination bacterium ZG1+ZG2)
Process 3:Cd
2++ add bacterium process (Cd
2++ bagasse immobilization combination bacterium ZG1+ZG2)
Process 4:Pb
2++ add bacterium process (Pb
2++ bagasse immobilization combination bacterium ZG1+ZG2)
Have 15 beakers under often kind of process, respectively at the 5th, 10,15,20, the residual quantity of 25d sampling and measuring phenanthrene, often kind of process 3 is parallel, keeps the water ratio of tested soil in each process to be about 15% in test.
After process terminates, measure the residual quantity of each process contaminated soil China and Philippines and calculate luxuriant and rich with fragrance degradation rate.For the soil of equal luxuriant and rich with fragrance pollution level, under different heavy metal ion exists, the degradation rate of soil China and Philippines changes as shown in Figure 4, heavy metal in soil ion Cd
2+and Pb
2existence bagasse immobilization combination bacterium ZG1+ZG2 phenanthrene pollutant soil repair is all had a certain impact, after process 25d, heavy metal free ion adds bacterium process, Cd
2++ add bacterium process and Pb
2++ add bacterium process to be respectively 95.85%, 80.56% and 85.02%, Cd
2+when existing, the degradation rate of soil China and Philippines is comparatively without Cd
2+degradation rate when existing decreases beyond 15%, even if but there is heavy metal ion Cd in soil
2+and Pb
2+, bagasse immobilization combination bacterium ZG1+ZG2 still can reach more than 80% to the degradation rate of soil China and Philippines.
The present embodiment illustrates that bagasse immobilization combination bacterium ZG1+ZG2 has good adaptive capacity to environment, for its application in complicated soil environment provides guarantee.
Claims (9)
1. a bagasse immobilization combination bacterium, it is characterized in that employing bagasse is the combination of carrier immobilized fungus and bacterium, wherein said fungi is trichoderma harziarum Pseudomonas, and described bacterium is Klebsiella.
2. a kind of bagasse immobilization combination bacterium according to claim 1, is characterized in that described trichoderma harziarum Pseudomonas fungi and Klebsiella bacteria are obtained through enrichment culture, separation and purification by the soil near petroleum chemical plant.
3. a kind of bagasse immobilization combination bacterium according to claim 2, is characterized in that described fungus and bacterium is prepared into fungi bacteria suspension and Bacteria suspension respectively through cultivation in advance when being inoculated in bagasse carrier.
4. a kind of bagasse immobilization combination bacterium according to claim 3, in 30 DEG C on the bagasse carrier that it is characterized in that described fungi bacteria suspension and Bacteria suspension to be inoculated in respectively the process of inorganic salt liquid nutrient solution, in the shaking table of rotating speed 150rpm, shaking culture is to biomass maximum and obtained immobilization combination bacterium.
5. a kind of bagasse immobilization combination bacterium according to claim 4, it is characterized in that described fungi bacteria suspension and Bacteria suspension are 1:1:2 ~ 6 (ml/ml/g) with the amount ratio of the bagasse processed, preferred fungi bacteria suspension and Bacteria suspension are 1:1:3 (ml/ml/g) with the amount ratio of the bagasse processed.
6. a kind of bagasse immobilization combination bacterium according to claim 3, is characterized in that described fungi bacteria suspension and Bacteria suspension are cultivated preparation by following steps and obtain:
Fungus and bacterium after purifying is inoculated in respectively in 50mL liquid nutrient medium, and pH is adjusted to 7.0, in 30 DEG C, on the shaking table of 150r/min, shaking culture 48h reaches maximum to the biomass of microorganism, gets the bacterium liquid of 4mL respectively in centrifuge tube, centrifugal 10min under 4000r/min, then supernatant liquor in centrifuge tube is poured out, supplementary phosphoric acid buffer continues centrifugal 10min, so repeats 3 times, and making bacteria suspension, to be placed in 4 DEG C of refrigerators for subsequent use.
7. a kind of bagasse immobilization combination bacterium according to claim 5, is characterized in that the step preparing immobilization combination bacterium is:
Take bagasse carrier, add after clean oven dry shreds in Erlenmeyer flask, aseptic inorganic salt liquid nutrient solution is added after sterilising treatment, add fungi bacteria suspension and Bacteria suspension more respectively, in 30 DEG C after mixing sealing, rotating speed be 150rpm shaking table in shaking culture 2 days namely obtain bagasse immobilization combination bacterium to microbes biomass is maximum, wherein said inorganic salt liquid nutrient solution and the amount ratio of bagasse carrier are 10:3 (mL/g).
8. the application of bagasse immobilization combination bacterium in soil Phenanthrene pollution amelioration according to any one of a claim 1-7.
9. the application of a kind of bagasse immobilization combination bacterium according to claim 8 in soil Phenanthrene pollution amelioration, is characterized in that described bagasse immobilization combination bacterium consumption is with the contaminated soil in its 10 times of weight of bagasse carrier 1 weight part process.
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Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106047364A (en) * | 2016-06-17 | 2016-10-26 | 战锡林 | Polycyclic aromatic hydrocarbons-contaminated soil remediation agent |
| CN106278468A (en) * | 2016-08-17 | 2017-01-04 | 山东省科学院生态研究所 | A kind of preparation method of biological organic fertilizer |
| CN106434614A (en) * | 2016-10-19 | 2017-02-22 | 华南农业大学 | Bagasse immobilized bacterium and application thereof to sulfamethoxazole pollution remediation of farmland soil |
| CN108977431A (en) * | 2018-06-25 | 2018-12-11 | 华南农业大学 | Bagasse immobilization microorganism Aspergillus aculeatus and its application in quinolone antibiotics pollution amelioration |
| CN110078208A (en) * | 2019-04-30 | 2019-08-02 | 广东省生物工程研究所（广州甘蔗糖业研究所） | A kind of immobilized active sludge and its preparation method and application |
| CN110153177A (en) * | 2019-03-08 | 2019-08-23 | 中国科学院广州地球化学研究所 | A method of polycyclic aromatic hydrocarbon pollution is repaired using fungi |
| CN111718867A (en) * | 2020-05-29 | 2020-09-29 | 北京理工大学 | Petroleum aromatic hydrocarbon degrading strain PB3 for producing biosurfactant and application thereof |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN106047364A (en) * | 2016-06-17 | 2016-10-26 | 战锡林 | Polycyclic aromatic hydrocarbons-contaminated soil remediation agent |
| CN106278468A (en) * | 2016-08-17 | 2017-01-04 | 山东省科学院生态研究所 | A kind of preparation method of biological organic fertilizer |
| CN106278468B (en) * | 2016-08-17 | 2019-11-29 | 山东省科学院生态研究所 | A kind of preparation method of biological organic fertilizer |
| CN106434614A (en) * | 2016-10-19 | 2017-02-22 | 华南农业大学 | Bagasse immobilized bacterium and application thereof to sulfamethoxazole pollution remediation of farmland soil |
| CN108977431A (en) * | 2018-06-25 | 2018-12-11 | 华南农业大学 | Bagasse immobilization microorganism Aspergillus aculeatus and its application in quinolone antibiotics pollution amelioration |
| CN110153177A (en) * | 2019-03-08 | 2019-08-23 | 中国科学院广州地球化学研究所 | A method of polycyclic aromatic hydrocarbon pollution is repaired using fungi |
| CN110153177B (en) * | 2019-03-08 | 2020-07-31 | 中国科学院广州地球化学研究所 | Method for repairing polycyclic aromatic hydrocarbon polluted soil by using fungi |
| CN110078208A (en) * | 2019-04-30 | 2019-08-02 | 广东省生物工程研究所（广州甘蔗糖业研究所） | A kind of immobilized active sludge and its preparation method and application |
| CN111718867A (en) * | 2020-05-29 | 2020-09-29 | 北京理工大学 | Petroleum aromatic hydrocarbon degrading strain PB3 for producing biosurfactant and application thereof |
| CN111718867B (en) * | 2020-05-29 | 2022-03-18 | 北京理工大学 | Petroleum aromatic hydrocarbon degrading strain PB3 for producing biosurfactant and application thereof |
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