CN102876616B - Pseudomonas aeruginosa for denitrification and phosphorous removal under different dissolved oxygen conditions and application of pseudomonas aeruginosa - Google Patents
Pseudomonas aeruginosa for denitrification and phosphorous removal under different dissolved oxygen conditions and application of pseudomonas aeruginosa Download PDFInfo
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- CN102876616B CN102876616B CN 201210395333 CN201210395333A CN102876616B CN 102876616 B CN102876616 B CN 102876616B CN 201210395333 CN201210395333 CN 201210395333 CN 201210395333 A CN201210395333 A CN 201210395333A CN 102876616 B CN102876616 B CN 102876616B
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Abstract
The invention relates to a pseudomonas aeruginosa with heterotrophic nitrification-aerobic denitrification and dephosphorization functions under different dissolved oxygen conditions and application of the pseudomonas aeruginosa. The bacterial strain can carry out growth metabolism by using organic carbon as a unique carbon source and ammonia nitrogen as a unique nitrogen source, the ammonia nitrogen can be respectively converted into gas products through the heterotrophic nitrification-aerobic denitrification effect under different dissolved oxygen conditions such as the oxygen deficit condition, the microaerobic condition and the aerobiotic condition, and the goal of denitrification is reached. The bacterial strain can also carry out growth metabolism by using nitrate nitrogen as a unique nitrogen source, the nitrate nitrogen can be converted into gaseous products through the dephosphorization effect under the oxygen deficit, microaerobic and aerobiotic conditions, and the goal of denitrification is reached. Meanwhile, the bacterial strain can also reach the goal of removing phosphate in water through converting inorganic phosphorus into self components under different dissolved oxygen conditions. Therefore, the bacterial strain can be used for synchronously removing elements such as carbon, nitrogen and phosphorus in the water under different dissolved oxygen conditions, and wide application prospects are realized in the wastewater treatment.
Description
Technical field
The present invention relates to a Pseudomonas aeruginosa strain (Pseudomonas aeruginosa) and the application in the wastewater biological denitrificaion dephosphorization thereof.This bacterial strain can be brought into play the heterotrophic nitrification-aerobic denitrification function under different dissolved oxygen concentration, and finishes in denitrogenation phosphatic removal, thereby reaches the purpose of waste water synchronous denitrification dephosphorizing.
Background technology
At present, a large amount of dischargings of industrial sewage and sanitary wastewater make China's nature and ecotope suffer havoc, have constituted great threat also for human life and health.The pollutent that exists in the waste water mainly includes materials such as machine thing, heavy metal and nitrogen phosphorus nutrition element, wherein a large amount of existence of nitrogen phosphoric can be brought out the eutrophication phenomenon in the water body, thereby cause serious ecological environment problem such as black smelly, the fishes and shrimps death of algal bloom, water body.The water environmental problems that is on the rise for reply, China also promulgates and has formulated stricter sewage drainage standard, " urban wastewater treatment firm pollutant emission standard " (GB18918-2002) is divided into first discharge standard A standard and B standard, and wherein the A standard has proposed higher emission request to total nitrogen and total phosphorus.Therefore, develop the hot issue that the denitrification dephosphorization technique of efficient energy-saving more becomes present research.
In the treatment process of present numerous denitrogenation of waste water dephosphorization, biological carbon and phosphorous removal is still the main means of removing nitrogen phosphoric in the waste water.Traditional biological denitrificaion theoretical basis is the nitrification and denitrification effect under the microbial process: at first, the organonitrogen in the waste water is decomposed under the heterotrophic bacterium effect and is converted into ammonia nitrogen, and this process is more easily carried out; Secondly, ammonia nitrogen in the waste water is converted into nitrite nitrogen under the effect of autotrophic microorganism nitrite bacteria, nitrite nitrogen further is converted into nitre nitrogen under the effect of nitrobacteria then, wherein the process of being finished jointly by nitrite bacteria and nitrobacteria that ammonia nitrogen is converted into nitre nitrogen is referred to as nitrification, this process need carries out in aerobic environment, the general requirement dissolved oxygen is higher than 2.0mg/L, and nitrification need be with the carbon source of inorganic carbon as the bacterial growth metabolism simultaneously; At last, be converted into nitrogen under the effect of nitre nitrogen in denitrifying bacterium in the waste water, finally finish the denitrogenation of waste water process, this process is called denitrification, and denitrifying bacterium need be with carbon source and the energy of organic carbon as thalli growth in this process, simultaneously carry out metabolism with nitre nitrogen and nitrite nitrogen as electron acceptor(EA) and just can finish denitrification, so this process can only carry out under anaerobic environment, the general requirement dissolved oxygen is lower than 0.5mg/L.Because nitrobacteria and the different demands of denitrifying bacterium to nutritive substance and dissolved oxygen, the biological denitrification process that gets up according to traditional biological denitrogenation theoretical developments can only separately carry out nitrifying process and denitrification process, thereby cause the wastewater biological denitrificaion technical matters tediously long, capital construction and working cost height, nitric efficiency is difficult to reach problems such as discharging standards.
In the wastewater biological dephosphorization process, phosphoric is mainly removed by the effect of polyP bacteria (PAOs), and detailed process can be divided into anaerobic phosphorus release and two stages of aerobic suction phosphorus.Under anaerobic, polyP bacteria absorbs the low-molecular-weight fatty acid (VFA) in the surrounding environment and changes into poly-(PHB) and is stored in the cell, required energy derive is in the glycolysis of endocellular sugar unit and the hydrolysis of poly-phosphate, and the phosphoric acid salt that poly-phosphate hydrolysis meanwhile produces is discharged into the extracellular; Under aerobic condition, polyP bacteria absorbs the phosphorus amount that exceeds self growth demand and is stored in the cell with the form of poly-phosphate from surrounding environment, and required energy derive is in the oxidative metabolism of PHB.Phosphoric in the waste water just is stored in the polyP bacteria body with the form of poly-phosphate like this, realizes the purpose of waste water dephosphorization then by the discharging of excess sludge.
By the mechanism of action of biological carbon and phosphorous removal as can be seen, for giving full play to the denitrogenation dephosphorizing function of microorganism, the different dissolved oxygen stages need be set in waste water treatment process, finish multistep biological processes such as anaerobic phosphorus release, anoxic denitrification, aerobic nitrification and suction phosphorus respectively.Simultaneously, in single Sludge System, itself just exists many contradiction biological denitrificaion and dephosphorization, nitrification needs long nitrobacteria of cycle in epoch, therefore need keep higher sludge age, and the cycle in epoch of polyP bacteria is shorter, and unique channel of dephosphorization is the discharging excess sludge, so the dephosphorization process need is kept short sludge age.And because denitrification process and dephosphorization process all need organic carbon source to keep the growth metabolism of denitrifying bacterium and polyP bacteria, and the organic composition in the waste water generally is very limited, and this just causes denitrogenation and dephosphorization process to exist competition to carbon source.There is the obvious suppression effect in the nitrate of nitrifying process generation simultaneously to the dispose procedure of polyP bacteria.The intrinsic contradictions that exist in these biological carbon and phosphorous removal processes become the bottleneck problem that improves denitrogenation dephosphorizing efficient in the sewage active sludge treatment process.
Problems at the existence of traditional biological denitrification dephosphorization technique, development in recent years many novel biological carbon and phosphorous removal theory and technology, more representational have short-cut nitrification and denitrification, Anammox, a denitrifying phosphorus removal technique etc., these new bio denitrification dephosphorization techniques have solved problems such as carbon source deficiency, energy expenditure be big to a certain extent, but still need anaerobic-anoxic-aerobic stage alternately, and reaction unit and operational condition are had higher requirement.
Heterotrophic nitrification-aerobic denitrification is a focus of the theoretical investigation of new bio denitrogenation in recent years.The heterotrophic nitrification phenomenon just was found in the nitrifying process at soil as far back as 20 beginnings of the century of 19 end of the centurys, the scholar was arranged subsequently successively about the report of heterotrophic nitrification phenomenon.The eighties in 20th century, Robertson etc. isolate aerobic denitrifying bacteria Thiosphaera pantotropha(first and now rename Paracoccus denitrificans Paracoccus denitrificans as in sulphur removal and denilrification system) and reported the existence that the denitrification enzyme is, the aerobic denitrification phenomenon becomes people's research focus immediately.Present isolated aerobic denitrifying bacteria mainly is present in Rhodopseudomonas, Alkaligenes, paracoccus and bacillus etc.Along with going deep into of research, it is found that some aerobic denitrifying bacterias have the ability of heterotrophic nitrification simultaneously, and the synchronous nitration and denitrification phenomenon under the aerobic condition and mechanism have been carried out deep research.Yet, under different dissolved oxygen conditions, all show strong heterotrophic nitrification-aerobic denitrification ability, and the bacterial strain that can have concurrently than strong phosphorus removal functional also rarely has report in denitrogenation.Bacterial strain among the present invention can be brought into play the heterotrophic nitrification-aerobic denitrification function under different dissolved oxygen conditions, concrete summary of the invention has been applied for patent of invention (application publication number CN101880645A); By to this bacterium further discovering in sewage treatment process, this bacterium shows stronger dephosphorization ability in the performance denitrogenation, all is significant on the biological carbon and phosphorous removal theory and practice is used.
Summary of the invention
The object of the present invention is to provide a strain under different dissolved oxygen conditions, all can bring into play the Pseudomonas aeruginosa of denitrogenation dephosphorizing function (Pseudomonas aeruginosa) and use, well solved the nitration denitrification that exists in traditional denitrification dephosphorization technique and release phosphorus and inhale the problem that phosphorus can't organic unity.
Bacterial strain provided by the invention has following feature: Gram-negative, thalline are shaft-like, do not form bud big gun and capsule; After nutrient agar is cultivated 2d, form the circular bacterium colony of 2-3mm, white, flat, smooth surface produces the yellow-green colour pigment; Its 16S rRNA has as the nuclear former times acid sequence shown in sequence is represented, sequence length is 1055bp.
According to strain morphology feature physiological and biochemical property and 16S rRNA gene sequence characteristic, identify that this bacterial strain is Pseudomonas aeruginosa (Pseudomonas aeruginosa).
Pseudomonas aeruginosa provided by the invention (Pseudomonas aeruginosa) is stored in China Committee for Culture Collection of Microorganisms common micro-organisms center (being called for short CGMCC) on January 21st, 2010, the depositary institution address is No. 3, Yard 1, BeiChen xi Road, Chaoyang District, Beijing City, Institute of Microorganism, Academia Sinica, postcode is 100101, and preserving number is CGMCC No.3602.
As the above-mentioned Pseudomonas aeruginosa that provides (Pseudomonas aeruginosa) bacterial strain CGMCC No.3602, it is characterized in that: this bacterial strain can utilize organic carbon to be sole carbon source, ammonia nitrogen or nitre nitrogen are that only nitrogen source carries out growth metabolism and brings into play the heterotrophic nitrification-aerobic denitrification effect, thereby nitrogen is removed.
As the above-mentioned Pseudomonas aeruginosa that provides (Pseudomonas aeruginosa) bacterial strain CGMCC No.3602, it is characterized in that: this bacterial strain can utilize organic carbon to be sole carbon source, ammonia nitrogen or nitre nitrogen are that only nitrogen source is removed the phosphoric acid salt in the water when carrying out the effect of growth metabolism performance heterotrophic nitrification-aerobic denitrification, thereby reach the purpose of nutritive elements such as nitrogen phosphorus in the synchronous removal water.
As the above-mentioned Pseudomonas aeruginosa that provides (Pseudomonas aeruginosa) bacterial strain CGMCC No.3602 in Application of Sewage, it is characterized in that: this bacterial strain can be at pH value 7-9, temperature 25-35 ℃, performance synchronous denitrification dephosphorizing function under anaerobism, anoxic, little different dissolved oxygen concentration such as aerobic and aerobic.
Application as the above-mentioned Pseudomonas aeruginosa that provides (Pseudomonas aeruginosa) bacterial strain CGMCC No.3602, it is characterized in that: this bacterial strain under anaerobic can be removed nitric nitrogen and the phosphoric acid salt in the water simultaneously, and anaerobic condition refers to the situation of cultivating in nitrogen environment.This bacterial strain is up to 99.83% to the clearance of nitre nitrogen under the anaerobic condition, and the highest clearance to phosphorus reaches 47.96% simultaneously.
Application as the above-mentioned Pseudomonas aeruginosa that provides (Pseudomonas aeruginosa) bacterial strain CGMCC No.3602, it is characterized in that: this bacterial strain can utilize ammonia nitrogen or nitric nitrogen to be only nitrogen source under anoxia condition, remove the phosphoric acid salt in the water and reach the purpose of dephosphorization, anoxia condition refers to the situation that places the biochemical incubator of 30-50rpm to cultivate substratum.
Application as the above-mentioned Pseudomonas aeruginosa that provides (Pseudomonas aeruginosa) bacterial strain CGMCC No.3602, it is characterized in that: this bacterial strain under anoxia condition is that nitrogenous source is when carrying out growth metabolism with the ammonia nitrogen, the highest clearance to ammonia nitrogen reaches 89.86%, removal speed is 1.73mgN/Lh, the phosphoric clearance is reached 99.71%, and removal speed is 0.29mgP/Lh.
Application as the above-mentioned Pseudomonas aeruginosa that provides (Pseudomonas aeruginosa) bacterial strain CGMCC No.3602, it is characterized in that: this bacterial strain is that nitrogenous source is when carrying out growth metabolism with nitre nitrogen under anoxia condition, be that the clearance of the phosphorus of 99.78%, 24h reaches 51.46% to the highest clearance of nitre nitrogen.
As the application of the above-mentioned Pseudomonas aeruginosa that provides (Pseudomonas aeruginosa) bacterial strain CGMCC No.3602, it is characterized in that: this bacterial strain reaches 98.66% to ammonia nitrogen removal frank under little aerobic condition, and removal speed is 2.03mgN/Lh; The phosphoric acid salt clearance is reached 100%.Under little aerobic condition, the nitre nitrogen removal efficiency is reached 95.22%, be 76.41% to the phosphate concn clearance, removal speed is 0.25mgP/Lh, and little aerobic condition refers to the situation that places the biochemical incubator of 70-80rpm to cultivate substratum, and dissolved oxygen is at 1.8-2.5mg/L.
As the application of the above-mentioned Pseudomonas aeruginosa that provides (Pseudomonas aeruginosa) bacterial strain CGMCC No.3602, it is characterized in that: this bacterial strain reaches 98.66% to ammonia nitrogen removal frank under aerobic condition, and removal speed is 2.03mgN/Lh; The phosphoric acid salt clearance is reached 100%.Under little aerobic condition, the nitre nitrogen removal efficiency is reached 95.22%, be 76.41% to the phosphate concn clearance, removal speed is 0.25mgP/Lh, and little aerobic condition refers to the situation that places the biochemical incubator of 70-80rpm to cultivate substratum, and dissolved oxygen is at 1.8-2.5mg/L.
As the application of the above-mentioned Pseudomonas aeruginosa that provides (Pseudomonas aeruginosa) bacterial strain CGMCC No.3602, it is characterized in that: this bacterial strain reaches 99.23% to ammonia nitrogen removal frank under aerobic condition, and removal speed is 7.39mgN/Lh; The phosphoric acid salt clearance is reached 87.11%, and removal speed is 1.07mgP/Lh.This bacterial strain is being that nitrogenous source reaches 73.91% to the nitre nitrogen removal efficiency when carrying out growth metabolism with nitre nitrogen, and removal speed is 3.04mgN/Lh; The phosphoric acid salt clearance is reached 99.55%, and removal speed is 0.64mgP/Lh.Aerobic condition refers to the situation that places the biochemical incubator of 140-180rpm to cultivate substratum, and dissolved oxygen is at 4.0-5.0mg/L.
Pseudomonas aeruginosa of the present invention (Pseudomonas aeruginosa) bacterial strain is used in denitrification dephosphorization technique has following advantage:
(1) has very strong heterotrophic nitrification-aerobic denitrification function according to this bacterial strain, can utilize this bacterium that the nitrogen in the water is removed in a stage fully, solved nitrification and denitrification in the traditional biological denitrification process distribute in time and space can not be unified problem, shorten technical process, saved capital cost.
(2) this bacterial strain is finished the nitrification and denitrification process a stage, and the basicity that denitrification process produces can remedy the acidity that produces in the nitrifying process, thereby does not need to add in addition acid-alkali accommodation pH value, thereby has saved running cost greatly.
(3) this bacterium can also effectively be removed the phosphoric in the water in the performance biological denitrificaion, has well solved sludge age disunity in denitrogenation and the dephosphorization process and carbon source is required to exist the problem of competition, has good economy and environmental benefit.
(4) this bacterium all can be brought into play the denitrogenation dephosphorizing function under different dissolved oxygen conditions, can utilize the microbial inoculum of this bacterium preparation to carry out biological reinforced to existing traditional biological denitrification dephosphorization technique, improve nitrogen phosphorus effluent quality, the cost of investment of having saved the technology upgrading to greatest extent simultaneously.
(5) the present invention is applicable to the multiple denitrogenation dephosphorizing processing that contains organic wastewater, has broad application prospects and good social benefit.
Description of drawings
Accompanying drawing 1 Pseudomonas aeruginosa is under anaerobic to nitre nitrogen and phosphatic degradation curve;
Accompanying drawing 2 Pseudomonas aeruginosas under anoxia condition to ammonia nitrogen and phosphatic degradation curve;
Accompanying drawing 3 Pseudomonas aeruginosas under anoxia condition to nitre nitrogen and phosphatic degradation curve;
Accompanying drawing 4 Pseudomonas aeruginosas under little aerobic condition to ammonia nitrogen and phosphatic degradation curve;
Accompanying drawing 5 Pseudomonas aeruginosas under little aerobic condition to nitre nitrogen and phosphatic degradation curve;
Accompanying drawing 6 Pseudomonas aeruginosas under aerobic condition to ammonia nitrogen and phosphatic degradation curve;
Accompanying drawing 7 Pseudomonas aeruginosas under aerobic condition to nitre nitrogen and phosphatic degradation curve.
Embodiment
Below in conjunction with accompanying drawing and embodiment the present invention is described in further detail, but the invention is not restricted to following examples.
Among the following embodiment, the determination and analysis method of all contaminations adopts potassium bichromate-sealed digestion method to measure with reference to " water and effluent monitoring analytical procedure " (the 4th edition, China Environmental Science Press, 2002): COD; OD
600Adopt ultraviolet spectrophotometer to measure, as reference, measure the absorbancy of bacteria suspension with nonvaccinated substratum at 600nm wavelength place; PH adopts accurate pH meter to measure; (YSI550A USA) measures by portable dissolved oxygen meter for temperature and dissolved oxygen.(SmartChem200 USA) measures by full-automatic nitrogen phosphorus analyser for ammonia nitrogen, nitrite nitrogen, nitre nitrogen and phosphoric acid salt phosphorus concentration.
Embodiment 1: Pseudomonas aeruginosa denitrification and dephosphorization ability under anaerobic measured
Bacterial strain (be stored in China Committee for Culture Collection of Microorganisms common micro-organisms center (being called for short CGMCC) on January 21st, 2010, preserving number is CGMCC No.3602, down together) with 20 ℃ of glycerine preservations of ﹣.(every L contains 5.0g Soduxin, 0.607gNaNO to be inoculated in sterilized 100mL substratum
3, 0.110g K
2HPO
43H
2O, 0.10g MgSO
47H
2O, 0.006g FeSO
47H
2O, 0.02g CaCl
22H
2O, (every L contains 3.5g EDTA, 2gZnSO to 0.10mL trace element mother liquor
47H
2O, 1g CuSO
45H
2O, 1g MnSO
47H
2O, 1g Na
2MoO
42H
2O, 1g H
3BO
3, 0.2gCoCl
26H
2O), pH 7.0 ~ 7.5) the 250mL Erlenmeyer flask in, place 30 ℃, shaking culture 20 ~ 24h in the 150rpm shaking table makes thalli growth to the logarithm later stage, bacteria suspension is used for inoculation.
Get in the 100mL anaerobism bottle that the above-mentioned bacteria suspension of 5mL joins the sterilized 100mL of containing test media (composition is with above-mentioned pre-culture medium), three parallel laboratory tests be set simultaneously, and with the substratum of not inoculating bacteria suspension as blank.Utilize vacuum pump to vacuumize then, charge into high-purity nitrogen again, repeat to vacuumize and inflated with nitrogen operation three times, at last the anaerobism bottle is placed 30 ℃, shaking culture in the 150rpm shaking table.
In culturing process, the substratum that takes a morsel at set intervals, wherein a part is directly used in and measures thalline optical density(OD) OD
600, rest part is centrifugal 5min under 8000rpm, gets supernatant liquor and measures indexs such as nitrogen nitrogen, nitrite nitrogen, nitre nitrogen, phosphorus, COD and pH.
Experimental result is seen accompanying drawing 1.As shown in the figure, Pseudomonas aeruginosa is in logarithmic phase at 0 ~ 30h, and cell density increases fast, and the nitre nitrogen concentration is quick downtrending simultaneously, drops to 0.15mg/L by initial 92.38mg/L, and clearance reaches 99.83%, and removal speed is 3.07mgN/Lh; Phosphate concn drops to Schwellenwert 8.25mg/L at 14h, clearance is 47.96%, removal speed is 0.54mgP/Lh, the trend that more significantly rebounds has appearred in phosphate concn subsequently, may be to cause owing to dissolving institute has appearred in thalline after release or thalline death have under anaerobic appearred in the phosphorus that early growth period absorbs; After cultivating 30h, Pseudomonas aeruginosa grows into stationary phase and decline phase, and it is stable that nitre nitrogen concentration and phosphate concn all reach.
The result can illustrate by experiment, Pseudomonas aeruginosa under anaerobic can carry out denitrification efficiently, the clearance of cultivating the nitre nitrogen of 30h reaches 99.83%, almost there is not simultaneously the accumulation of nitrite nitrogen, in denitrification process, Pseudomonas aeruginosa also has the dephosphorization ability, and the clearance of cultivating the phosphorus of 14h reaches 47.96%.
Embodiment 2: the nitrated and dephosphorization ability of Pseudomonas aeruginosa under anoxia condition measured
(every L contains 5.0g Soduxin, 0.607gNaNO to the inoculation that 20 ℃ of glycerine of ﹣ are preserved in sterilized 100mL substratum
3, 0.110g K
2HPO
43H
2O, 0.10g MgSO
47H
2O, 0.006g FeSO
47H
2O, 0.02g CaCl
22H
2O, (every L contains 3.5g EDTA to 0.10mL trace element mother liquor, 2g ZnSO
47H
2O, 1gCuSO
45H
2O, 1gMnSO
47H
2O, 1gNa
2MoO
42H
2O, 1gH
3BO
3, 0.2g CoCl
26H
2O), pH 7.0 ~ 7.5) the 250mL Erlenmeyer flask in, place 30 ℃, shaking culture 20 ~ 24h in the 150rpm shaking table makes thalli growth to the logarithm later stage, bacteria suspension is used for inoculation.
Get in the 250mL Erlenmeyer flask that the above-mentioned bacteria suspension of 5mL joins the sterilized 100mL of containing test media (composition is with above-mentioned substratum), place 30 ℃, the interior shaking culture of 40rpm shaking table, experimental group arranges three parallel laboratory tests, and with the substratum of not inoculating bacteria suspension as blank.
In culturing process, the substratum that takes a morsel at set intervals, wherein a part is directly used in and measures thalline optical density(OD) OD
600, rest part is centrifugal 5min under 8000rpm, gets supernatant liquor and measures indexs such as nitrogen nitrogen, nitrite nitrogen, nitre nitrogen, phosphorus, COD and pH.
Experimental result is seen accompanying drawing 2.As shown in the figure, Pseudomonas aeruginosa is in logarithmic phase at 0 ~ 30h, and cell density increases fast; In the 36h that cultivates, ammonia nitrogen concentration and phosphate concn be downtrending once always, and wherein ammonia nitrogen concentration drops to 7.02mg/L by initial 69.22mg/L, and clearance reaches 89.86%, and removal speed is 1.73mgN/Lh; Phosphate concn drops to 0.03mg/L by initial 10.47mg/L, and clearance reaches 99.71%, and removal speed is 0.29mgP/Lh.
The result as can be seen by experiment, Pseudomonas aeruginosa can be removed ammonia nitrogen efficiently under anoxia condition, cultivate the ammonia-N removal rate of 36h and reach 89.86%, there are not nitrite nitrogen and nitre nitrogen and accumulation simultaneously, when removing ammonia nitrogen, Pseudomonas aeruginosa also has very strong dephosphorization ability, and the clearance of the phosphorus of 36h reaches 99.71%, illustrates that Pseudomonas aeruginosa has stronger phosphorus removal functional concurrently in the effect of performance heterotrophic nitrification.
Embodiment 3: denitrification and the dephosphorization ability of Pseudomonas aeruginosa under anoxia condition measured
(every L contains 5.0g Soduxin, 0.607gNaNO to the inoculation that 20 ℃ of glycerine of ﹣ are preserved in sterilized 100mL substratum
3, 0.110g K
2HPO
43H
2O, 0.10g MgSO
47H
2O, 0.006g FeSO
47H
2O, 0.02g CaCl
22H
2O, (every L contains 3.5g EDTA to 0.10mL trace element mother liquor, 2g ZnSO
47H
2O, 1gCuSO
45H
2O, 1gMnSO
47H
2O, 1gNa
2MoO
42H
2O, 1gH
3BO
3, 0.2g CoCl
26H
2O), pH 7.0 ~ 7.5) the 250mL Erlenmeyer flask in, place 30 ℃, shaking culture 20 ~ 24h in the 150rpm shaking table makes thalli growth to the logarithm later stage, bacteria suspension is used for inoculation.
Get in the 250mL Erlenmeyer flask that the above-mentioned bacteria suspension of 5mL joins the sterilized 100mL of containing test media (composition is with above-mentioned pre-culture medium), place 30 ℃, the interior shaking culture of 40rpm shaking table, experimental group arranges three parallel laboratory tests, and with the substratum of not inoculating bacteria suspension as blank.
In culturing process, the substratum that takes a morsel at set intervals, wherein a part is directly used in and measures thalline optical density(OD) OD
600, rest part is centrifugal 5min under 8000rpm, gets supernatant liquor and measures indexs such as nitrogen nitrogen, nitrite nitrogen, nitre nitrogen, phosphorus, COD and pH.
Experimental result is seen accompanying drawing 3.As shown in the figure, Pseudomonas aeruginosa is in logarithmic phase at 0 ~ 32h, and cell density increases fast, enters the stable growth phase subsequently; At preceding 12h, hysteresis phenomenon appears in the degraded of nitre nitrogen, removes comparatively slowly, and the nitre nitrogen concentration presents quick downtrending afterwards, drops to 0.21mg/L when 40h, and clearance reaches 99.78%, and removal speed is 2.47mgN/Lh; Phosphate concn drops to Schwellenwert 7.46mg/L at 24h, clearance is 51.46%, removal speed is 0.33mgP/Lh, the trend that more significantly rebounds has appearred in phosphate concn subsequently, may be because the dissolving of organophosphorus has appearred in the phosphorus that thalline absorbs at early growth period after having occurred release or thalline death under the anoxia condition cause; At 40 ~ 46h, diauxie has appearred in thalline, and phosphate concn has obtained degraded again.
The result can illustrate by experiment, Pseudomonas aeruginosa can carry out denitrification efficiently under anoxia condition, the clearance of cultivating the nitre nitrogen of 40h reaches 99.78%, almost there is not simultaneously the accumulation of nitrite nitrogen, in denitrification process, Pseudomonas aeruginosa also has stronger dephosphorization ability, and the clearance of cultivating the phosphorus of 24h reaches 51.46%.
Embodiment 4: the nitrated and dephosphorization ability of Pseudomonas aeruginosa under little aerobic condition measured
(every L contains 5.0g Soduxin, 0.607gNaNO to the inoculation that 20 ℃ of glycerine of ﹣ are preserved in sterilized 100mL substratum
3, 0.110g K
2HPO
43H
2O, 0.10g MgSO
47H
2O, 0.006g FeSO
47H
2O, 0.02g CaCl
22H
2O, (every L contains 3.5g EDTA to 0.10mL trace element mother liquor, 2g ZnSO
47H
2O, 1gCuSO
45H
2O, 1gMnSO
47H
2O, 1gNa
2MoO
42H
2O, 1gH
3BO
3, 0.2g CoCl
26H
2O), pH 7.0 ~ 7.5) the 250mL Erlenmeyer flask in, place 30 ℃, shaking culture 20 ~ 24h in the 150rpm shaking table makes thalli growth to the logarithm later stage, bacteria suspension is used for inoculation.
Get in the 250mL Erlenmeyer flask that the above-mentioned bacteria suspension of 5mL joins the sterilized 100mL of containing test media (composition is with above-mentioned substratum), place 30 ℃, the interior shaking culture of 75rpm shaking table, experimental group arranges three parallel laboratory tests, and with the substratum of not inoculating bacteria suspension as blank.
In culturing process, the substratum that takes a morsel at set intervals, wherein a part is directly used in and measures thalline optical density(OD) OD
600, rest part is centrifugal 5min under 8000rpm, gets supernatant liquor and measures indexs such as nitrogen nitrogen, nitrite nitrogen, nitre nitrogen, phosphorus, COD and pH.
Experimental result is seen accompanying drawing 4.As shown in the figure, Pseudomonas aeruginosa is in logarithmic phase at 0 ~ 42h, and cell density increases fast; Meanwhile, ammonia nitrogen concentration and phosphate concn are synchronous downtrending, and wherein ammonia nitrogen concentration drops to 1.16mg/L by initial 86.25mg/L, and clearance reaches 98.66%, and removal speed is 2.03mgN/Lh; Phosphate concn drops to 0.00mg/L by initial 13.57mg/L, and clearance reaches 100%, and removal speed is 0.32mgP/Lh.
The result as can be seen by experiment, Pseudomonas aeruginosa can be removed ammonia nitrogen efficiently under little aerobic condition, cultivate the ammonia-N removal rate of 42h and reach 98.66%, there are not nitrite nitrogen and nitre nitrogen and accumulation simultaneously, when removing ammonia nitrogen, Pseudomonas aeruginosa also has very strong dephosphorization ability, and 42h can remove phosphorus fully, illustrates that Pseudomonas aeruginosa has very strong phosphorus removal functional concurrently in the effect of performance heterotrophic nitrification.
Embodiment 5: denitrification and the dephosphorization ability of Pseudomonas aeruginosa under little aerobic condition measured
(every L contains 5.0g Soduxin, 0.607gNaNO to the inoculation that 20 ℃ of glycerine of ﹣ are preserved in sterilized 100mL substratum
3, 0.110g K
2HPO
43H
2O, 0.10g MgSO
47H
2O, 0.006g FeSO
47H
2O, 0.02g CaCl
22H
2O, (every L contains 3.5g EDTA to 0.10mL trace element mother liquor, 2g ZnSO
47H
2O, 1gCuSO
45H
2O, 1gMnSO
47H
2O, 1gNa
2MoO
42H
2O, 1gH
3BO
3, 0.2g CoCl
26H
2O), pH 7.0 ~ 7.5) the 250mL Erlenmeyer flask in, place 30 ℃, shaking culture 20 ~ 24h in the 150rpm shaking table makes thalli growth to the logarithm later stage, bacteria suspension is used for inoculation.
Get in the 250mL Erlenmeyer flask that the above-mentioned bacteria suspension of 5mL joins the sterilized 100mL of containing test media (composition is with above-mentioned pre-culture medium), place 30 ℃, the interior shaking culture of 75rpm shaking table, experimental group arranges three parallel laboratory tests, and with the substratum of not inoculating bacteria suspension as blank.
In culturing process, the substratum that takes a morsel at set intervals, wherein a part is directly used in and measures thalline optical density(OD) OD
600, rest part is centrifugal 5min under 8000rpm, gets supernatant liquor and measures indexs such as nitrogen nitrogen, nitrite nitrogen, nitre nitrogen, phosphorus, COD and pH.
Experimental result is seen accompanying drawing 5.As shown in the figure, Pseudomonas aeruginosa is in logarithmic phase at 0 ~ 42h, and cell density increases fast, subsequently because the consumption of nutritive substance and the accumulation thalline of harmful meta-bolites enter decline phase; Nitre nitrogen and phosphatic clearance all reach the highest at cultivation 48h, and wherein the nitre nitrogen concentration drops to 5.12mg/L by initial 107.14mg/L, and clearance reaches 95.22%, and removal speed is 2.13mgN/Lh; Phosphate concn drops to 3.65mg/L by initial 15.46mg/L, and clearance reaches 76.41%, and removal speed is 0.25mgP/Lh, and nitre nitrogen concentration and phosphate concn all keep stable subsequently.
The result can illustrate by experiment, Pseudomonas aeruginosa can carry out denitrification efficiently under little aerobic condition, the clearance of cultivating the nitre nitrogen of 42h reaches 95.22%, almost there is not simultaneously the accumulation of nitrite nitrogen, in denitrification process, Pseudomonas aeruginosa also has stronger dephosphorization ability, and the clearance of cultivating the phosphorus of 42h reaches 76.41%.
Embodiment 6: the nitrated and dephosphorization ability of Pseudomonas aeruginosa under aerobic condition measured
(every L contains 5.0g Soduxin, 0.607gNaNO to the inoculation that 20 ℃ of glycerine of ﹣ are preserved in sterilized 100mL substratum
3, 0.110g K
2HPO
43H
2O, 0.10g MgSO
47H
2O, 0.006g FeSO
47H
2O, 0.02g CaCl
22H
2O, (every L contains 3.5g EDTA to 0.10mL trace element mother liquor, 2g ZnSO
47H
2O, 1gCuSO
45H
2O, 1gMnSO
47H
2O, 1gNa
2MoO
42H
2O, 1gH
3BO
3, 0.2g CoCl
26H
2O), pH 7.0 ~ 7.5) the 250mL Erlenmeyer flask in, place 30 ℃, shaking culture 20 ~ 24h in the 150rpm shaking table makes thalli growth to the logarithm later stage, bacteria suspension is used for inoculation.
Get in the 250mL Erlenmeyer flask that the above-mentioned bacteria suspension of 5mL joins the sterilized 100mL of containing test media (composition is with above-mentioned substratum), place 30 ℃, the interior shaking culture of 150rpm shaking table, experimental group arranges three parallel laboratory tests, and with the substratum of not inoculating bacteria suspension as blank.
In culturing process, the substratum that takes a morsel at set intervals, wherein a part is directly used in and measures thalline optical density(OD) OD
600, rest part is centrifugal 5min under 8000rpm, gets supernatant liquor and measures indexs such as nitrogen nitrogen, nitrite nitrogen, nitre nitrogen, phosphorus, COD and pH.
Experimental result is seen accompanying drawing 6.As shown in the figure, Pseudomonas aeruginosa is in logarithmic phase at 0 ~ 9h, and cell density increases fast, and thalline enters stationary phase subsequently; It can also be seen that ammonia nitrogen and phosphoric acid salt are removed when cultivating 12h substantially fully, wherein ammonia nitrogen concentration drops to 0.68mg/L by initial 89.33mg/L, and clearance reaches 99.23%, and removal speed is 7.39mgN/Lh; Phosphate concn drops to 1.90mg/L by initial 14.70mg/L, and clearance reaches 87.11%, and removal speed is 1.07mgP/Lh.
The result as can be seen by experiment, Pseudomonas aeruginosa can be removed ammonia nitrogen efficiently under aerobic condition, cultivate the ammonia-N removal rate of 12h and can reach 99.23%, there are not nitrite nitrogen and nitre nitrogen and accumulation simultaneously, when removing ammonia nitrogen, Pseudomonas aeruginosa also has very strong dephosphorization ability, and the clearance of cultivating the phosphorus of 12h can reach 87.11%, illustrates that Pseudomonas aeruginosa has very strong phosphorus removal functional concurrently in the effect of performance heterotrophic nitrification.
Embodiment 7: denitrification and the dephosphorization ability of Pseudomonas aeruginosa under aerobic condition measured
(every L contains 5.0g Soduxin, 0.607gNaNO to the inoculation that 20 ℃ of glycerine of ﹣ are preserved in sterilized 100mL substratum
3, 0.110g K
2HPO
43H
2O, 0.10g MgSO
47H
2O, 0.006g FeSO
47H
2O, 0.02g CaCl
22H
2O, (every L contains 3.5g EDTA to 0.10mL trace element mother liquor, 2g ZnSO
47H
2O, 1gCuSO
45H
2O, 1gMnSO
47H
2O, 1gNa
2MoO
42H
2O, 1gH
3BO
3, 0.2g CoCl
26H
2O), pH 7.0 ~ 7.5) the 250mL Erlenmeyer flask in, place 30 ℃, shaking culture 20 ~ 24h in the 150rpm shaking table makes thalli growth to the logarithm later stage, bacteria suspension is used for inoculation.
Get in the 250mL Erlenmeyer flask that the above-mentioned bacteria suspension of 5mL joins the sterilized 100mL of containing test media (composition is with above-mentioned pre-culture medium), place 30 ℃, the interior shaking culture of 150rpm shaking table, experimental group arranges three parallel laboratory tests, and with the substratum of not inoculating bacteria suspension as blank.
In culturing process, the substratum that takes a morsel at set intervals, wherein a part is directly used in and measures thalline optical density(OD) OD
600, rest part is centrifugal 5min under 8000rpm, gets supernatant liquor and measures indexs such as nitrogen nitrogen, nitrite nitrogen, nitre nitrogen, phosphorus, COD and pH.
Experimental result is seen accompanying drawing 7.As shown in the figure, Pseudomonas aeruginosa is in logarithmic phase at 0 ~ 24h, and cell density increases fast; Meanwhile, nitre nitrogen and phosphate concn are degraded fast, and wherein the nitre nitrogen concentration drops to 25.72mg/L by initial 98.59mg/L, and clearance reaches 73.91%, and removal speed is 3.04mgN/Lh; Phosphate concn drops to 0.07mg/L by initial 15.52mg/L, and clearance reaches 99.55%, and removal speed is 0.64mgP/Lh.
The result can illustrate by experiment, Pseudomonas aeruginosa can carry out denitrification under aerobic condition, the clearance of cultivating the nitre nitrogen of 24h reaches 73.91%, almost there is not simultaneously the accumulation of nitrite nitrogen, in denitrification process, Pseudomonas aeruginosa also has stronger dephosphorization ability, and the clearance of cultivating the phosphorus of 24h reaches 99.55%.
Claims (5)
1. a strain has the application of Pseudomonas aeruginosa (Pseudomonas aeruginosa) in wastewater treatment of heterotrophic nitrification-aerobic denitrification and phosphorus removal functional concurrently, it is characterized in that: described Pseudomonas aeruginosa was stored in China Committee for Culture Collection of Microorganisms common micro-organisms center (being called for short CGMCC) on January 21st, 2010, preserving number is No.3602, described application refers to: at pH value 7-9, remove elements such as carbon in the water, nitrogen, phosphorus under temperature 25-35 ℃ of condition simultaneously.
2. the application of the described Pseudomonas aeruginosa of claim 1 (Pseudomonas aeruginosa) in wastewater treatment, it is characterized in that: under anaerobic, can remove nitric nitrogen and phosphoric acid salt in the water simultaneously, anaerobic condition refers to the situation of cultivating in nitrogen environment.
3. the application of the described Pseudomonas aeruginosa of claim 1 (Pseudomonas aeruginosa) in wastewater treatment, it is characterized in that: under anoxia condition, utilize ammonia nitrogen or nitric nitrogen to be only nitrogen source, remove the phosphoric acid salt in the water and reach the purpose of dephosphorization, anoxia condition refers to the situation that places the biochemical incubator of 30-50rpm to cultivate substratum.
4. the application of the described Pseudomonas aeruginosa of claim 1 (Pseudomonas aeruginosa) in wastewater treatment, it is characterized in that: under little aerobic condition, utilize ammonia nitrogen or nitric nitrogen to be only nitrogen source, reach the purpose of removing nitrogen phosphorus in the water simultaneously, little aerobic condition refers to the situation that places the biochemical incubator of 70-80rpm to cultivate substratum, and dissolved oxygen is at 1.8-2.5mg/L.
5. the application of the described Pseudomonas aeruginosa of claim 1 (Pseudomonas aeruginosa) in wastewater treatment, it is characterized in that: under aerobic condition, utilize ammonia nitrogen or nitric nitrogen to be only nitrogen source, phosphoric reaches the purpose of denitrogenation dephosphorizing simultaneously in the body thereby simultaneously inorganic phosphorus in the water is converted into, aerobic condition refers to the situation that places the biochemical incubator of 140-180rpm to cultivate substratum, and dissolved oxygen is at 4.0-5.0mg/L.
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| CN103304040B (en) * | 2013-05-23 | 2014-07-09 | 东北电力大学 | Method for treating nitrogen-containing waste water by utilizing carbon fiber felt-immobilized pseudomonas aeruginosa |
| CN107177530B (en) * | 2017-06-03 | 2020-02-18 | 农业农村部环境保护科研监测所 | Novel efficient domestic sewage denitrifying bacterium and application thereof |
| CN109055252A (en) * | 2017-11-13 | 2018-12-21 | 鼎正新兴生物技术（天津）有限公司 | A kind of heterotrophic nitrification-aerobic denitrification complex microorganism preparations and preparation method thereof |
| CN110038423A (en) * | 2019-05-22 | 2019-07-23 | 天津渤化永利化工股份有限公司 | The system and method for pseudomonas aeruginosa degradation butyl octanol unit VOCs exhaust gas |
| CN112299566A (en) * | 2019-07-24 | 2021-02-02 | 天津农学院 | New method for removing nitrogen and phosphorus |
| CN110468066B (en) * | 2019-07-29 | 2021-09-17 | 中成进出口股份有限公司 | Aerobic denitrifying strain and application thereof |
| CN112175863B (en) * | 2020-09-15 | 2022-05-13 | 重庆理工大学 | Preparation method and application of complex microbial inoculant for removing nitrogen-containing pollutants in high-salinity wastewater |
| CN114480159B (en) * | 2021-11-23 | 2024-03-29 | 北京工业大学 | Synchronous heterotrophic nitrification aerobic denitrification dephosphorization bacterium and application thereof |
| CN115322936B (en) * | 2022-09-21 | 2023-10-27 | 北京大学 | Aerobic denitrifying bacterium with sulfonamide antibiotic degradation capability and application thereof |
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| CN101857847A (en) * | 2010-05-06 | 2010-10-13 | 合肥工业大学 | Pseudomonas aeruginosa strain separating, purifying and domesticating method and use |
| CN101880645A (en) * | 2010-06-04 | 2010-11-10 | 北京大学 | Verdigris pseudomonas aeruginosa strain denitrified under different dissolved oxygen conditions and application thereof |
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| CN101857847A (en) * | 2010-05-06 | 2010-10-13 | 合肥工业大学 | Pseudomonas aeruginosa strain separating, purifying and domesticating method and use |
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