CN104726376A - Continuous large-scale culture method for high-concentration salt-resistant nitrifying bacteria - Google Patents
Continuous large-scale culture method for high-concentration salt-resistant nitrifying bacteria Download PDFInfo
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Abstract
The invention discloses a continuous large-scale culture method for high-concentration salt-resistant nitrifying bacteria, and relates to the field of environmental biotechnology. The method comprises an enrichment-separation phase and a culture-separation phase, wherein directional enrichment is carried out by adding an enrichment culture solution, large-scale culture is carried out by adding a growth nutrient solution, precipitation separation is carried out, and the high-concentration salt-resistant nitrifying bacteria are finally obtained. The salt-resistant nitrifying bacteria prepared by the culture method disclosed by the invention is high in concentration, high in pollutant removal efficiency, low in energy consumption, and suitable for biotreatment for high-salt ammonia-nitrogen wastewater.
Description
Technical field
The present invention relates to field of environmental biotechnology, particularly relate to a kind of high density salt tolerant nitrobacteria large-scaled culture method of continous way.
Background technology
Along with the promulgation of urban wastewater treatment firm emission standard (GB18918-2002), total nitrogen includes discharge index in first.On December 15th, 2011; State Council prints and distributes " national environmental protection " 12 " planning "; first ammonia nitrogen is classified as the restrictive Con trolling index of the main water pollutions discharge in the whole nation, explicitly pointing out to ammonia nitrogen total emission volumn in 2015 is 2,380,000 tons, cuts down 10% than the same period in 2010.High slat-containing wastewater refers generally to the waste water that total salinity (with NaCl content meter) is at least 1%, is extensively present in the industries such as chemical industry, pharmacy, process hides, food, oil recovery, marine products processing.Due to the ammonia nitrogen (about 1000 ~ 5000mg/L) of high salt industrial waste water generally containing high density, if can not qualified discharge, then non-compliant high-salt wastewater can polluted surface, soil, coastal and river mouth etc., causes various environmental problem and the ecological problems such as such as eutrophication.Therefore, realize high slat-containing wastewater biological denitrificaion how effectively, economically and be treated as Science and engineering difficulties urgently to be resolved hurrily.
Materialization denitrogenation mainly contains chemical neutralization method, chemical precipitation method, break point chlorination method, selective ion exchange method, air and stripping denitrogenation etc.Relative to materialization denitrogenation, and biological method reasonably utilizes the physiological function of nitrifier and denitrifying bacteria, and the nitrogen of the various forms in sewage finally can be converted into gaseous nitrogen, the possibility of the secondary pollution of generation is less, processing cost is lower.Thus bio-denitrification technology has more actual use value.
Halophilic microorganism is the extreme microorganism that a class lives in hypersaline environment, is extensively present in the hypersaline environments such as saltern, salt lake, soil.According to salinity scope (1 ~ 30%), halophilic microorganism is divided into Facultative Halophiles, slight halophilic bacterium, Halophilic Bacterium and Natrinema altunense sp.These halophilic microorganisms form unique height and ooze the ability of surviving in environment in long-term evolutionary process, have very special physiological structure and metabolic mechanism.The aspects such as the character of the stability of the cytolemma of halophilic microorganism, cell wall structure composition and functional ingredient, reaction kinetics, enzyme system, pathways metabolism and information transmission, protein nucleic acid component and conformation have specificity for adapting to hypersaline environment.These Mechanisms of Salt Resistances ensure that halophilic microorganism carries out metabolism and growth in hypersaline environment.Therefore only take the mode progressively improving ammonia nitrogen concentration, selective elimination does not possess the salt tolerant bacterium of nitrification, and orienting enriching, to the salt tolerant nitrobacteria of high density, then carries out pilot scale culture, can be applied to engineering.
In existing culture technique; China patent document CN 101240253 A discloses a kind of method adopting active sludge intermittent type enrichment nitrifier; with ammonia nitrogen, small-scale inorganic salt and damping fluid for nutrient solution; its NH_3-N treating scope≤1200mg/L; the method does not add organic substance, and nitrobacteria ultimate density changes too greatly, when processing high-concentration ammonia nitrogenous wastewater; there is the possibility of degraded in mud, is therefore not suitable for pilot scale culture.China patent document CN1017092278 discloses a kind of sequence batch (that adopts and intakes in conjunction with the large-scaled culture method of flocculation sediment enrichment nitrococcus; although the method is enriched to high density nitrococcus; but most nitrifier is eliminated; cause water outlet nitrite very high, final outflow water total nitrogen is very high.China patent document CN1354786A discloses the cultural method of a kind of active sludge middle and high concentration nitrobacteria, dirty mud Xi seed sludge is urinated with downflow sludge and dung, with sludge dewatering filtrate or nitrated disengaging liquid for nutrient solution, process ammonia nitrogen scope 100 ~ 300mg/L, exceeding this concentration can produce restraining effect to thalline.
Summary of the invention
Technical problem to be solved by this invention is for the deficiencies in the prior art, provides a kind of method design reasonable, can cultivate the high density salt tolerant nitrobacteria large-scaled culture method of the continous way obtaining the bacterium product that concentration is high, contaminant removal efficiency is high.
Technical problem to be solved by this invention is realized by following technical scheme.The present invention is the large-scaled culture method of a kind of high density salt tolerant nitrobacteria of continous way, be characterized in: the method comprises Rich Internet Applications stage and cultivation-separation phase, take to add enrichment culture liquid to carry out orienting enriching, add growth nutrient solution and carry out pilot scale culture, the final nitrobacteria obtaining high density; Its concrete steps are as follows:
(1) the Rich Internet Applications stage: prepare initial ammonia nitrogen mass concentration 50 ~ 150mg/L, salinity massfraction in the enrichment nutritive medium of NaCl≤10% with water, flow into incubator by volume pump; The alkali lye of preparation massfraction 1% ~ 5%, flows into incubator by volume pump; Get mud sample and 5% ~ 20% be once seeded to incubator by volume, carry out the orienting enriching of salt tolerant nitrobacteria; The mud mixed liquid that enrichment obtains is separated by precipitate and separate device, and supernatant liquor is discharged from precipitate and separate device top, and the salt tolerant nitrobacteria that enrichment obtains is back to incubator;
(2) cultivation-separation phase: after the Rich Internet Applications stage terminates, prepares initial ammonia nitrogen mass concentration 400mg/L, salinity massfraction in the growth nutrient solution of NaCl≤10% with water, flows into incubator by volume pump; The alkali lye of preparation massfraction 1% ~ 5%, flows into incubator by volume pump; After adding growth nutrient solution, carry out the pilot scale culture of nitrobacteria; Cultivate the mud mixed liquid obtained to be separated by precipitate and separate device, a bottom part collects the salt tolerant nitrobacteria of high density, and a part of salt tolerant nitrobacteria mud mixed liquid is back to incubator in addition, and supernatant liquor is discharged from precipitate and separate device top;
The condition of Rich Internet Applications stage and cultivation-separation phase is as follows: temperature is 10 DEG C ~ 35 DEG C, and pH is 6.0 ~ 10.0; DO is 1.5 ~ 7.5 mg/L.
The large-scaled culture method of the high density salt tolerant nitrobacteria of a kind of continous way of the present invention, its further preferred technical scheme or technical characteristic as follows:
1, enrichment nutritive medium of the present invention can adopt any one disclosed in prior art to go for the enrichment nutritive medium of salt tolerant nitrobacteria of the present invention, composition and the consumption of preferred enrichment nutritive medium are: the ammonium salt of 0.5 ~ 10g/L, the KH of 0.5 ~ 5.0 g/L
2pO
4or K
2hPO
4, the MgCl of 0.5 ~ 5.0 g/L
2, the Na of 0.5 ~ 5.0 g/L
2cO
3, the NaCl of massfraction≤10%, liquid microelement;
Described growth nutrient solution can adopt any one disclosed in prior art to go for the growth nutrient solution of salt tolerant nitrobacteria of the present invention, and the composition of preferred growth nutrient solution and consumption are: 0.5 ~ 10g/Lr ammonium salt, the KH of 0.5 ~ 10g/L
2pO
4or K
2hPO
4, the MgCl of 0.5 ~ 10g/L
2, the Na of 0.5 ~ 10g/L
2cO
3, the glucose of 0.5 ~ 10g/L or methyl alcohol, the NaCl of massfraction≤10%, liquid microelement;
Described liquid microelement can adopt any one disclosed in prior art to go for the liquid microelement of salt tolerant nitrobacteria of the present invention, and composition and the consumption thereof of preferred liquid microelement are as follows: the CoCl of 50 ~ 150mg/L
26H
2the CaCl of O, 50 ~ 150mg/L
2, the CuSO of 10 ~ 50mg/L
45H
2the ZnSO of O, 50 ~ 150mg/L
47H
2the FeSO of O, 50 ~ 150mg/L
47H
2the MnSO of O, 50 ~ 150mg/L
42H
2o.
In the present invention, composition and the consumption of further preferred enrichment nutritive medium are: ammonium salt (3 ~ 6g/L), KH
2pO
4or K
2hPO
4(2 ~ 3g/L), MgCl
2(2 ~ 3 g/L), Na
2cO
3(2 ~ 3g/L), NaCl(massfraction≤10%), liquid microelement.Composition and the consumption of further preferred growth nutrient solution are: ammonium salt (3 ~ 6g/L), KH
2pO
4or K
2hPO
4(3 ~ 6g/L), MgCl
2(3 ~ 6g/L), Na
2cO
3(3 ~ 6g/L), glucose or methyl alcohol (3 ~ 6g/L), NaCl(massfraction≤10%), liquid microelement.The component of preferred liquid microelement and consumption thereof are: CoCl
26H
2o(80 ~ 120mg/L), CaCl
2(80 ~ 120mg/L), CuSO
45H
2o(25 ~ 35mg/L), ZnSO
47H
2o(80 ~ 120mg/L), FeSO
47H
2o(80 ~ 120mg/L), MnSO
42H
2o(80 ~ 120mg/L).
2, enrichment nutritive medium of the present invention, the ammonium salt described in growth nutrient solution can select the conventional ammonium salt being applicable to salt tolerant nitrobacteria, and preferred ammonium salt is selected from one or more of ammonium sulfate, urea or ammonium chloride.When selecting the ammonium salt of mixing, its blending ratio can be chosen arbitrarily on demand.
3, according to step of the present invention (1) described enriching method, preferably every 1 ~ 2d is 1 enrichment cycle, and the enrichment cycle is preferably 3 ~ 5; According to the cultural method described in step of the present invention (2), every 2d ~ 3d is preferably 1 culture cycle, and culture cycle is preferably 5 ~ 8.
4, in the inventive method, the Rich Internet Applications stage preferably adopts the method progressively improving ammonia nitrogen concentration, and wherein ammonia nitrogen starting point concentration is 50 ~ 150mg/L, and ultimate density is 500 ~ 1000mg/L; Concrete grammar is as follows: after adding enrichment nutritive medium at every turn, when ammonia nitrogen concentration is down to below 5mg/L, improves the ammonia nitrogen concentration in enrichment nutritive medium, increase rate 20 ~ 100 mg/L; The dosage of suitable raising alkali lye, to guarantee the pH value environment required for nitrobacteria growth.In aforesaid method, ammonia nitrogen starting point concentration more preferably 80 ~ 120mg/L, ultimate density is 700 ~ 800mg/L more preferably.
5, in the inventive method, cultivation-separation phase preferably adopts stable ammonia nitrogen concentration, carries out concentration range correction simultaneously; Wherein stable ammonia nitrogen concentration is 400mg/L, and concentration correction scope is 300 ~ 500mg/L; Concrete grammar is as follows: add ammonia nitrogen concentration in grown cultures liquid is 400mg/L at every turn, as ammonia nitrogen concentration≤5mg/L, suitably improves ammonia nitrogen concentration and corrects, increase rate 10 ~ 100mg/L; As ammonia nitrogen concentration >=15 mg/L, suitably reduce ammonia nitrogen concentration and correct, reduce amplitude 10 ~ 100mg/L.
6, in the inventive method, the salt tolerant nitrobacteria mud mixed liquid volume fraction ratio that cultivation-separation phase is back to incubator bottom precipitate and separate device preferably controls 10 ~ 40%, preferably controls 20 ~ 30% further, most preferably controls 25%.
7, in the inventive method, the ratio of the backflow flow velocity of cultivation-separation phase from precipitate and separate device to incubator preferably controls at 0.1 ~ 1:1, controls at 0.4 ~ 0.7:1 further, most preferably controls at 0.5:1.
8, in the inventive method, Rich Internet Applications stage and cultivation-separation phase alkali lye used can adopt any one disclosed in prior art to go for the alkali lye of salt tolerant nitrobacteria of the present invention, and preferred alkali lye is NaOH, Na
2cO
3, NaHCO
3and CaCO
3in solution one or more, when selecting mixed alkali liquor, its blending ratio can select arbitrary proportion on demand.
9, the inventive method, can add appropriate active carbon powder, to improve its effect within Rich Internet Applications stage and cultivation-separation phase incubator.
10, Rich Internet Applications stage and cultivation-separation phase condition are temperature is 10 DEG C ~ 35 DEG C; PH is 6.0 ~ 10.0; DO is 1.5 ~ 7.5 mg/L.
In the inventive method, can saltern be taken from containing mud sample, also can take from other sources as the hypersaline environment such as salt lake, soil.
The population succession phenomenon of the inventive method in the Rich Internet Applications stage according to Microbial Communities in Activated Sludge group, adopt and progressively improve ammonia nitrogen mass concentration, make the miscellaneous bacterias such as the protozoon in mud, metazoan, fungi and other heterotrophic bacteriums be suppressed at Different growth phases, progressively eliminate " non-nitrobacteria ", salt tolerant nitrobacteria is made to become dominant microflora, final nitrobacteria can the ammonia nitrogen waste water of enduring high-concentration, has higher ammonia nitrogen removal frank.At cultivation-separation phase, maintain certain ammonia nitrogen mass concentration, the miscellaneous bacterias such as protozoon with this understanding, metazoan, fungi and other heterotrophic bacteriums are suppressed, and salt tolerant nitrobacteria becomes dominant microflora, by adding certain glucose, methyl alcohol or other carbon sources, salt tolerant nitrobacteria is grown fast, collects and obtain high density salt tolerant nitrobacteria.
That uses due to nitrobacteria cultivation stage mostly is cheap carbon source, makes its large-scale production become possibility.Nitrobacteria is widely used in the biological treatment of various Industrial Wastewater Treatment, can also be applied in various municipal wastewater treatment plant and start fast and problem emergent management.
The beneficial effect that the present invention compared with prior art has is as follows:
(1) to cultivate the salt tolerant nitrobacteria concentration of preparation high in the present invention, and active good, cultural method used is applicable to large-scale production.
(2) there is stronger ammonia nitrogen removal ability, the ammonia nitrogen waste water of lower concentration can not only be processed, ideal to the ammonia nitrogen removal effect of high density, have a good application prospect.
(3) the salt tolerant nitrobacteria prepared by the present invention itself, to environment toxicological harmless effect, effectively can reduce residue mud generation, and reduce the use of the pharmaceutical chemicalss such as flocculation agent.Therefore the cost of biochemical treatment ammonia nitrogen waste water and energy consumption lower.
Accompanying drawing explanation
Fig. 1 is the changing conditions of Rich Internet Applications stage water-in and water-out ammonia nitrogen and ammonia nitrogen removal frank;
Fig. 2 is the changing conditions of cultivation-separation phase water-in and water-out ammonia nitrogen, MLSS;
Fig. 3 is cultural method schematic diagram of the present invention;
Fig. 4 is salt tolerant nitrobacteria process urea production enterprise wastewater test-results.
Embodiment
Below in conjunction with drawings and Examples, further describe the solution of the present invention and effect.Should be appreciated that, concrete case study on implementation described below only for better explaining the present invention, but does not limit the present invention.
Embodiment 1, the large-scaled culture method of a kind of high density salt tolerant nitrobacteria of continous way, the method comprises Rich Internet Applications stage and cultivation-separation phase, take to add enrichment culture liquid to carry out orienting enriching, add growth nutrient solution and carry out pilot scale culture, the final nitrobacteria obtaining high density; Its concrete steps are as follows:
(1) the Rich Internet Applications stage: prepare initial ammonia nitrogen mass concentration 50mg/L, salinity massfraction in the enrichment nutritive medium of NaCl≤10% with water, flow into incubator by volume pump; The alkali lye of preparation massfraction 1%, flows into incubator by volume pump; Get mud sample and 5% be once seeded to incubator by volume, carry out the orienting enriching of salt tolerant nitrobacteria; The mud mixed liquid that enrichment obtains is separated by precipitate and separate device, and supernatant liquor is discharged from precipitate and separate device top, and the salt tolerant nitrobacteria that enrichment obtains is back to incubator;
(2) cultivation-separation phase: after the Rich Internet Applications stage terminates, prepares initial ammonia nitrogen mass concentration 400mg/L, salinity massfraction in the growth nutrient solution of NaCl≤10% with water, flows into incubator by volume pump; The alkali lye of preparation massfraction 1%, flows into incubator by volume pump; After adding growth nutrient solution, carry out the pilot scale culture of nitrobacteria; Cultivate the mud mixed liquid obtained to be separated by precipitate and separate device, a bottom part collects the salt tolerant nitrobacteria of high density, and a part of salt tolerant nitrobacteria mud mixed liquid is back to incubator in addition, and supernatant liquor is discharged from precipitate and separate device top;
The condition of Rich Internet Applications stage and cultivation-separation phase is as follows: temperature is 10 DEG C ~ 35 DEG C, and pH is 6.0 ~ 10.0; DO is 1.5 ~ 7.5 mg/L.
Embodiment 2, the large-scaled culture method of a kind of high density salt tolerant nitrobacteria of continous way, the method comprises Rich Internet Applications stage and cultivation-separation phase, take to add enrichment culture liquid to carry out orienting enriching, add growth nutrient solution and carry out pilot scale culture, the final nitrobacteria obtaining high density; Its concrete steps are as follows:
(1) the Rich Internet Applications stage: prepare initial ammonia nitrogen mass concentration 150mg/L, salinity massfraction in the enrichment nutritive medium of NaCl≤10% with water, flow into incubator by volume pump; The alkali lye of preparation massfraction 5%, flows into incubator by volume pump; Get mud sample and 20% be once seeded to incubator by volume, carry out the orienting enriching of salt tolerant nitrobacteria; The mud mixed liquid that enrichment obtains is separated by precipitate and separate device, and supernatant liquor is discharged from precipitate and separate device top, and the salt tolerant nitrobacteria that enrichment obtains is back to incubator;
(2) cultivation-separation phase: after the Rich Internet Applications stage terminates, prepares initial ammonia nitrogen mass concentration 400mg/L, salinity massfraction in the growth nutrient solution of NaCl≤10% with water, flows into incubator by volume pump; The alkali lye of preparation massfraction 5%, flows into incubator by volume pump; After adding growth nutrient solution, carry out the pilot scale culture of nitrobacteria; Cultivate the mud mixed liquid obtained to be separated by precipitate and separate device, a bottom part collects the salt tolerant nitrobacteria of high density, and a part of salt tolerant nitrobacteria mud mixed liquid is back to incubator in addition, and supernatant liquor is discharged from precipitate and separate device top;
The condition of Rich Internet Applications stage and cultivation-separation phase is as follows: temperature is 20 DEG C ~ 25 DEG C, and pH is 7.0 ~ 9.0; DO is 3 ~ 5 mg/L.
Embodiment 3, the large-scaled culture method of a kind of high density salt tolerant nitrobacteria of continous way, the method comprises Rich Internet Applications stage and cultivation-separation phase, take to add enrichment culture liquid to carry out orienting enriching, add growth nutrient solution and carry out pilot scale culture, the final nitrobacteria obtaining high density; Its concrete steps are as follows:
(1) the Rich Internet Applications stage: prepare initial ammonia nitrogen mass concentration 100mg/L, salinity massfraction in the enrichment nutritive medium of NaCl≤10% with water, flow into incubator by volume pump; The alkali lye of preparation massfraction 3%, flows into incubator by volume pump; Get mud sample and 10% be once seeded to incubator by volume, carry out the orienting enriching of salt tolerant nitrobacteria; The mud mixed liquid that enrichment obtains is separated by precipitate and separate device, and supernatant liquor is discharged from precipitate and separate device top, and the salt tolerant nitrobacteria that enrichment obtains is back to incubator;
(2) cultivation-separation phase: after the Rich Internet Applications stage terminates, prepares initial ammonia nitrogen mass concentration 400mg/L, salinity massfraction in the growth nutrient solution of NaCl≤10% with water, flows into incubator by volume pump; The alkali lye of preparation massfraction 3%, flows into incubator by volume pump; After adding growth nutrient solution, carry out the pilot scale culture of nitrobacteria; Cultivate the mud mixed liquid obtained to be separated by precipitate and separate device, a bottom part collects the salt tolerant nitrobacteria of high density, and a part of salt tolerant nitrobacteria mud mixed liquid is back to incubator in addition, and supernatant liquor is discharged from precipitate and separate device top;
The condition of Rich Internet Applications stage and cultivation-separation phase is as follows: temperature is 15 DEG C ~ 30 DEG C, and pH is 7.5 ~ 9.5; DO is 2 ~ 6 mg/L.
Embodiment 4, in the large-scaled culture method of the high density salt tolerant nitrobacteria of a kind of continous way described in embodiment 1 or 2 or 3: composition and the consumption of described enrichment nutritive medium are: the ammonium salt of 0.5 ~ 10g/L, the KH of 0.5 ~ 5.0 g/L
2pO
4or K
2hPO
4, the MgCl of 0.5 ~ 5.0 g/L
2, the Na of 0.5 ~ 5.0 g/L
2cO
3, the NaCl of massfraction≤10%, liquid microelement;
The composition of described growth nutrient solution and consumption are: 0.5 ~ 10g/Lr ammonium salt, the KH of 0.5 ~ 10g/L
2pO
4or K
2hPO
4, the MgCl of 0.5 ~ 10g/L
2, the Na of 0.5 ~ 10g/L
2cO
3, the glucose of 0.5 ~ 10g/L or methyl alcohol, the NaCl of massfraction≤10%, liquid microelement;
Composition and the consumption thereof of described liquid microelement are as follows: the CoCl of 50 ~ 150mg/L
26H
2the CaCl of O, 50 ~ 150mg/L
2, the CuSO of 10 ~ 50mg/L
45H
2the ZnSO of O, 50 ~ 150mg/L
47H
2the FeSO of O, 50 ~ 150mg/L
47H
2the MnSO of O, 50 ~ 150mg/L
42H
2o.
Embodiment 5, in the large-scaled culture method of the high density salt tolerant nitrobacteria of a kind of continous way of embodiment 1-4 described in any one: described ammonium salt is selected from one or more of ammonium sulfate, urea or ammonium chloride.
Embodiment 6, in the large-scaled culture method of the high density salt tolerant nitrobacteria of a kind of continous way of embodiment 1-5 described in any one: according to step (1) described enriching method, every 1d is 1 enrichment cycle, and the enrichment cycle is 3; According to the cultural method described in step (2), every 2d is 1 culture cycle, and culture cycle is 5.
Embodiment 7, in the large-scaled culture method of the high density salt tolerant nitrobacteria of a kind of continous way of embodiment 1-5 described in any one: according to step (1) described enriching method, every 2d is 1 enrichment cycle, and the enrichment cycle is 5; According to the cultural method described in step (2), every 3d is 1 culture cycle, and culture cycle is 8.
Embodiment 8, in the large-scaled culture method of the high density salt tolerant nitrobacteria of a kind of continous way of embodiment 1-7 described in any one: the Rich Internet Applications stage adopts the method progressively improving ammonia nitrogen concentration, wherein ammonia nitrogen starting point concentration is 50 ~ 150mg/L, and ultimate density is 500 ~ 1000mg/L; Concrete grammar is as follows: after adding enrichment nutritive medium at every turn, when ammonia nitrogen concentration is down to below 5mg/L, improves the ammonia nitrogen concentration in enrichment nutritive medium, increase rate 20 ~ 100 mg/L; The dosage of suitable raising alkali lye, to guarantee the pH value environment required for nitrobacteria growth.
Embodiment 9, in the large-scaled culture method of the high density salt tolerant nitrobacteria of a kind of continous way of embodiment 1-8 described in any one: cultivation-separation phase adopts stable ammonia nitrogen concentration, carries out concentration range correction simultaneously; Wherein stable ammonia nitrogen concentration is 400mg/L, and concentration correction scope is 300 ~ 500mg/L; Concrete grammar is as follows: add ammonia nitrogen concentration in grown cultures liquid is 400mg/L at every turn, as ammonia nitrogen concentration≤5mg/L, suitably improves ammonia nitrogen concentration and corrects, increase rate 10 ~ 100mg/L; As ammonia nitrogen concentration >=15 mg/L, suitably reduce ammonia nitrogen concentration and correct, reduce amplitude 10 ~ 100mg/L.
Embodiment 10, in the large-scaled culture method of the high density salt tolerant nitrobacteria of a kind of continous way of embodiment 1-9 described in any one: the salt tolerant nitrobacteria mud mixed liquid volume fraction ratio that cultivation-separation phase is back to incubator bottom precipitate and separate device controls 10% ~ 40%.
Embodiment 11, in the large-scaled culture method of the high density salt tolerant nitrobacteria of a kind of continous way of embodiment 1-9 described in any one: the salt tolerant nitrobacteria mud mixed liquid volume fraction ratio that cultivation-separation phase is back to incubator bottom precipitate and separate device controls 20 ~ 30%.
Embodiment 12, in the large-scaled culture method of the high density salt tolerant nitrobacteria of a kind of continous way of embodiment 1-11 described in any one: the ratio of the backflow flow velocity of cultivation-separation phase from precipitate and separate device to incubator controls at 0.1 ~ 1:1.
Embodiment 13, in the large-scaled culture method of the high density salt tolerant nitrobacteria of a kind of continous way of embodiment 1-11 described in any one: the ratio of the backflow flow velocity of cultivation-separation phase from precipitate and separate device to incubator controls at 0.4 ~ 0.6:1.
Embodiment 14, in the large-scaled culture method of the high density salt tolerant nitrobacteria of a kind of continous way of embodiment 1-13 described in any one: Rich Internet Applications stage and cultivation-separation phase alkali lye used are NaOH, Na
2cO
3, NaHCO
3and CaCO
3in solution one or more.
Embodiment 15, in the large-scaled culture method of the high density salt tolerant nitrobacteria of a kind of continous way of embodiment 1-14 described in any one: add appropriate active carbon powder within Rich Internet Applications stage and cultivation-separation phase incubator.
Embodiment 16, with reference to Fig. 1-3, the large-scaled culture method experiment one of a kind of high density salt tolerant nitrobacteria of continous way, mud sample takes from Lianyungang of Jiangsu saltern:
1. the Rich Internet Applications stage: the enrichment nutritive medium preparing initial ammonia nitrogen mass concentration 150mg/L adds nutritive medium storage tank, and enrichment nutritive medium flows into incubator by volume pump.The alkali lye of preparation massfraction 1% ~ 5% adds alkali liquor storage tank, flows into incubator by volume pump.Getting mud sample 30% is inoculated into incubator by volume, carry out the enrichment culture of salt tolerant nitrobacteria.Mud mixed liquid is separated by precipitate and separate device, and supernatant liquor is discharged from precipitate and separate device top, and the salt tolerant nitrobacteria of enrichment is back to incubator.
Wherein ammonium salt (0.5 ~ 10g/L), KH
2pO
4or K
2hPO
4(0.5 ~ 5.0 g/L), MgCl
2(0.5 ~ 5.0 g/L), Na
2cO
3(0.5 ~ 5.0 g/L), NaCl(massfraction≤10%), liquid microelement.The composition of liquid microelement is: CoCl
26H
2o(50 ~ 150mg/L), CaCl
2(50 ~ 150mg/L), CuSO
45H
2o(10 ~ 50mg/L), ZnSO
47H
2o(50 ~ 150mg/L), FeSO
47H
2o(50 ~ 150mg/L), MnSO
42H
2o(50 ~ 150mg/L).Period controls water temperature about 25.0 DEG C, and pH is 8.5, and dissolved oxygen is 3.0 mg/L.
According to aforesaid method, every 1d is 1 enrichment cycle, and the enrichment cycle is 3.
The whole enrichment culture stage adopts the method progressively improving ammonia nitrogen concentration to carry out enrichment, and wherein ammonia nitrogen starting point concentration is 100mg/L, and ultimate density is 900mg/L.Concrete grammar is as follows: after adding enrichment nutritive medium at every turn, when ammonia nitrogen concentration is down to below 5mg/L, improves the concentration of the ammonia nitrogen in enrichment nutritive medium, increase rate 50 ~ 100 mg/L.Suitable raising Na
2cO
3dosage, with guarantee nitrobacteria growth required for pH value environment.
2. cultivation-separation phase: after concentration stage terminates, prepares initial ammonia nitrogen mass concentration 400mg/, salinity massfraction≤10%(in NaCl) growth nutrient solution, flow into incubator by volume pump.The alkali lye of preparation massfraction 1% ~ 5%, flows into incubator by volume pump.After adding growth nutrient solution, carry out the pilot scale culture of nitrobacteria.Mud mixed liquid is separated by precipitate and separate device, the salt tolerant nitrobacteria of the high density of bottom collection 80%, and other 20% salt tolerant nitrobacteria is back to incubator, and supernatant liquor is discharged from precipitate and separate device top.
Wherein the composition of grown cultures liquid is: ammonium salt (0.5 ~ 10g/L), KH
2pO
4or K
2hPO
4(0.5 ~ 10g/L), MgCl
2(0.5 ~ 10g/L), Na
2cO
3(0.5 ~ 10g/L), glucose or methyl alcohol (0.5 ~ 10g/L), NaCl(massfraction≤10%), liquid microelement.The composition of liquid microelement is: CoCl
26H
2o(50 ~ 150mg/L), CaCl
2(50 ~ 150mg/L), CuSO
45H
2o(10 ~ 50mg/L), ZnSO
47H
2o(50 ~ 150mg/L), FeSO
47H
2o(50 ~ 150mg/L), MnSO
42H
2o(50 ~ 150mg/L).Period controls water temperature about 25.0 DEG C, and pH is 9.5, and dissolved oxygen is 4.5 mg/L.
According to the method described above, every 3d is 1 culture cycle, and culture cycle is 6.
Whole cultivation stage adopts stable ammonia nitrogen concentration, carries out concentration range correction simultaneously.Wherein stable ammonia nitrogen concentration is 400mg/L, and concentration correction scope is 300 ~ 500mg/L.Concrete grammar is as follows: after adding grown cultures liquid at every turn, and ammonia nitrogen concentration is 400mg/L, as ammonia nitrogen concentration≤5mg/L, improves ammonia nitrogen concentration and corrects, increase rate 10 ~ 100mg/L; As ammonia nitrogen concentration >=15 mg/L, reduce ammonia nitrogen concentration and correct, reduce amplitude 10 ~ 100mg/L.
Embodiment 17, with reference to Fig. 1-3, the large-scaled culture method experiment two of a kind of high density salt tolerant nitrobacteria of continous way, mud sample takes from Lianyungang of Jiangsu saltern:
1. the Rich Internet Applications stage: the enrichment nutritive medium preparing initial ammonia nitrogen mass concentration 100mg/L adds nutritive medium storage tank, and enrichment nutritive medium flows into incubator by volume pump.The alkali lye of preparation massfraction 1% ~ 5% adds alkali liquor storage tank, flows into incubator by volume pump.Getting mud sample 10% is inoculated into incubator by volume, carry out the enrichment culture of salt tolerant nitrobacteria.Mud mixed liquid is separated by precipitate and separate device, and supernatant liquor is discharged from precipitate and separate device top, and the salt tolerant nitrobacteria of enrichment is back to incubator.
Wherein ammonium salt (0.5 ~ 10g/L), KH
2pO
4or K
2hPO
4(0.5 ~ 5.0 g/L), MgCl
2(0.5 ~ 5.0 g/L), Na
2cO
3(0.5 ~ 5.0 g/L), NaCl(massfraction≤10%), liquid microelement.The composition of liquid microelement is: CoCl
26H
2o(50 ~ 150mg/L), CaCl
2(50 ~ 150mg/L), CuSO
45H
2o(10 ~ 50mg/L), ZnSO
47H
2o(50 ~ 150mg/L), FeSO
47H
2o(50 ~ 150mg/L), MnSO
42H
2o(50 ~ 150mg/L).Period controls water temperature about 25.0 DEG C, and pH is 8.5, and dissolved oxygen is 3.0 mg/L.
According to aforesaid method, every 1d is 1 enrichment cycle, and the enrichment cycle is 3.
The whole enrichment culture stage adopts the method progressively improving ammonia nitrogen concentration to carry out enrichment, and wherein ammonia nitrogen starting point concentration is 100mg/L, and ultimate density is 900mg/L.Concrete grammar is as follows: after adding enrichment nutritive medium at every turn, when ammonia nitrogen concentration is down to below 5mg/L, improves the concentration of the ammonia nitrogen in enrichment nutritive medium, increase rate 50 ~ 100 mg/L.Suitable raising Na
2cO
3dosage, with guarantee nitrobacteria growth required for pH value environment.
2. cultivation-separation phase: after concentration stage terminates, prepares initial ammonia nitrogen mass concentration 400mg/, salinity massfraction≤10%(in NaCl) growth nutrient solution, flow into incubator by volume pump.The alkali lye of preparation massfraction 1% ~ 5%, flows into incubator by volume pump.After adding growth nutrient solution, carry out the pilot scale culture of nitrobacteria.Mud mixed liquid is separated by precipitate and separate device, the salt tolerant nitrobacteria of the high density of bottom collection 80%, and other 20% salt tolerant nitrobacteria is back to incubator, and supernatant liquor is discharged from precipitate and separate device top.
Wherein the composition of grown cultures liquid is: ammonium salt (0.5 ~ 10g/L), KH
2pO
4or K
2hPO
4(0.5 ~ 10g/L), MgCl
2(0.5 ~ 10g/L), Na
2cO
3(0.5 ~ 10g/L), glucose or methyl alcohol (0.5 ~ 10g/L), NaCl(massfraction≤10%), liquid microelement.The composition of liquid microelement is: CoCl
26H
2o(50 ~ 150mg/L), CaCl
2(50 ~ 150mg/L), CuSO
45H
2o(10 ~ 50mg/L), ZnSO
47H
2o(50 ~ 150mg/L), FeSO
47H
2o(50 ~ 150mg/L), MnSO
42H
2o(50 ~ 150mg/L).Period controls water temperature about 25.0 DEG C, and pH is 9.5, and dissolved oxygen is 4.5 mg/L.
According to the method described above, every 3d is 1 culture cycle, and culture cycle is 6.
Whole cultivation stage adopts stable ammonia nitrogen concentration, carries out concentration range correction simultaneously.Wherein stable ammonia nitrogen concentration is 400mg/L, and concentration correction scope is 300 ~ 500mg/L.Concrete grammar is as follows: after adding grown cultures liquid at every turn, and ammonia nitrogen concentration is 400mg/L, as ammonia nitrogen concentration≤5mg/L, improves ammonia nitrogen concentration and corrects, increase rate 10 ~ 100mg/L; As ammonia nitrogen concentration >=15 mg/L, reduce ammonia nitrogen concentration and correct, reduce amplitude 10 ~ 100mg/L.
Embodiment 18, with reference to Fig. 1-3, the large-scaled culture method experiment three of a kind of high density salt tolerant nitrobacteria of continous way, mud sample takes from Lianyungang of Jiangsu saltern:
1. the Rich Internet Applications stage: the enrichment nutritive medium preparing initial ammonia nitrogen mass concentration 200mg/L adds nutritive medium storage tank, and enrichment nutritive medium flows into incubator by volume pump.The alkali lye of preparation massfraction 1% ~ 5% adds alkali liquor storage tank, flows into incubator by volume pump.Getting mud sample 40% is inoculated into incubator by volume, carry out the enrichment culture of salt tolerant nitrobacteria.Mud mixed liquid is separated by precipitate and separate device, and supernatant liquor is discharged from precipitate and separate device top, and the salt tolerant nitrobacteria of enrichment is back to incubator.
Wherein ammonium salt (0.5 ~ 10g/L), KH
2pO
4or K
2hPO
4(0.5 ~ 5.0 g/L), MgCl
2(0.5 ~ 5.0 g/L), Na
2cO
3(0.5 ~ 5.0 g/L), NaCl(massfraction≤10%), liquid microelement.The composition of liquid microelement is: CoCl
26H
2o(50 ~ 150mg/L), CaCl
2(50 ~ 150mg/L), CuSO
45H
2o(10 ~ 50mg/L), ZnSO
47H
2o(50 ~ 150mg/L), FeSO
47H
2o(50 ~ 150mg/L), MnSO
42H
2o(50 ~ 150mg/L).Period controls water temperature about 25.0 DEG C, and pH is 8.5, and dissolved oxygen is 3.0 mg/L.
According to aforesaid method, every 1d is 1 enrichment cycle, and the enrichment cycle is 3.
The whole enrichment culture stage adopts the method progressively improving ammonia nitrogen concentration to carry out enrichment, and wherein ammonia nitrogen starting point concentration is 100mg/L, and ultimate density is 900mg/L.Concrete grammar is as follows: after adding enrichment nutritive medium at every turn, when ammonia nitrogen concentration is down to below 5mg/L, improves the concentration of the ammonia nitrogen in enrichment nutritive medium, increase rate 50 ~ 100 mg/L.Suitable raising Na
2cO
3dosage, with guarantee nitrobacteria growth required for pH value environment.
2. cultivation-separation phase: after concentration stage terminates, prepares initial ammonia nitrogen mass concentration 400mg/, salinity massfraction≤10%(in NaCl) growth nutrient solution, flow into incubator by volume pump.The alkali lye of preparation massfraction 1% ~ 5%, flows into incubator by volume pump.After adding growth nutrient solution, carry out the pilot scale culture of nitrobacteria.Mud mixed liquid is separated by precipitate and separate device, the salt tolerant nitrobacteria of the high density of bottom collection 80%, and other 20% salt tolerant nitrobacteria is back to incubator, and supernatant liquor is discharged from precipitate and separate device top.
Wherein the composition of grown cultures liquid is: ammonium salt (0.5 ~ 10g/L), KH
2pO
4or K
2hPO
4(0.5 ~ 10g/L), MgCl
2(0.5 ~ 10g/L), Na
2cO
3(0.5 ~ 10g/L), glucose or methyl alcohol (0.5 ~ 10g/L), NaCl(massfraction≤10%), liquid microelement.The composition of liquid microelement is: CoCl
26H
2o(50 ~ 150mg/L), CaCl
2(50 ~ 150mg/L), CuSO
45H
2o(10 ~ 50mg/L), ZnSO
47H
2o(50 ~ 150mg/L), FeSO
47H
2o(50 ~ 150mg/L), MnSO
42H
2o(50 ~ 150mg/L).Period controls water temperature about 25.0 DEG C, and pH is 9.5, and dissolved oxygen is 4.5 mg/L.
According to the method described above, every 3d is 1 culture cycle, and culture cycle is 6.
Whole cultivation stage adopts stable ammonia nitrogen concentration, carries out concentration range correction simultaneously.Wherein stable ammonia nitrogen concentration is 400mg/L, and concentration correction scope is 300 ~ 500mg/L.Concrete grammar is as follows: after adding grown cultures liquid at every turn, and ammonia nitrogen concentration is 400mg/L, as ammonia nitrogen concentration≤5mg/L, improves ammonia nitrogen concentration and corrects, increase rate 10 ~ 100mg/L; As ammonia nitrogen concentration >=15 mg/L, reduce ammonia nitrogen concentration and correct, reduce amplitude 10 ~ 100mg/L.
Cultivate the removal effect experiment to simulation ammonia nitrogen waste water of the nitrobacteria that obtains by method described in embodiment 16-18, process and result as follows:
Adopt the bio-aeration pool of 10L, count at salinity massfraction 8%(NaCl) under condition under, preparing ammonia nitrogen mass concentration is respectively 440 ~ 462mg/L, 649 ~ 662 mg/L, the simulation ammonia nitrogen waste water of 838 ~ 882 mg/L, get 200mL embodiment 16 respectively, embodiment 17, the high density salt tolerant nitrobacteria that embodiment 18 is cultivated, inoculum size according to 20% is added in the bio-aeration pool of 10L respectively, control dissolved oxygen is 2.5mg/L ~ 6.5mg/L, temperature is about 28 DEG C, adding appropriate sodium bicarbonate makes pH maintain 7.0 ~ 8.5, sampling and measuring water outlet ammonia nitrogen concentration after aeration time 24h.Result shows that ammonia nitrogen removal frank reaches more than 99% after 24h process, reaches ideal process effect.In table 1.
Table 1. microbial inoculum treatment effect
Cultivate by method described in embodiment 16-18 the salt tolerant nitrobacteria obtained to test the removal effect of urea production enterprise wastewater, process and result as follows:
Urea waste water takes from Lianyun Harbour nitrogen fertilizer production enterprise wastewater, and wherein ammonia nitrogen mass concentration is 350 ~ 550 mg/L.Because in former water, ammonia nitrogen mass concentration is higher, therefore ordinary activated sludge is difficult to the ammonia nitrogen removed wherein, and water outlet ammonia nitrogen mass concentration is far above qualified discharge standard.Add certain industrial NaCl, make its salinity massfraction be 5%.
Urea waste water process adopts SBR technique, the salt tolerant nitrobacteria in Example 1, and according in the inoculum size access SBR device of volume ratio 20%, every day was undertaken by 1 cycle, and water inlet 15min, aeration 22h, sedimentation 30min, draining 15min, residue 1h is idle mixing time.Wherein Process operating parameters is as follows: dissolved oxygen is 2.5mg/L ~ 6.5mg/L, and temperature is about 28 ~ 32 DEG C, adds sodium bicarbonate and makes pH maintain 7.0 ~ 8.0.Continuum micromeehanics 14d, test achieves desirable effect.Average influent ammonium concentration is 448.5 mg/L, and average water outlet ammonia nitrogen concentration is 1.7mg/L, and average ammonia nitrogen removal frank reaches 99.6%.As shown in Figure 4.
Claims (10)
1. the large-scaled culture method of the high density salt tolerant nitrobacteria of a continous way, it is characterized in that: the method comprises Rich Internet Applications stage and cultivation-separation phase, take to add enrichment culture liquid to carry out orienting enriching, add growth nutrient solution and carry out pilot scale culture, the final nitrobacteria obtaining high density; Its concrete steps are as follows:
(1) the Rich Internet Applications stage: prepare initial ammonia nitrogen mass concentration 50 ~ 150mg/L, salinity massfraction in the enrichment nutritive medium of NaCl≤10% with water, flow into incubator by volume pump; The alkali lye of preparation massfraction 1% ~ 5%, flows into incubator by volume pump; Get mud sample and 5% ~ 20% be once seeded to incubator by volume, carry out the orienting enriching of salt tolerant nitrobacteria; The mud mixed liquid that enrichment obtains is separated by precipitate and separate device, and supernatant liquor is discharged from precipitate and separate device top, and the salt tolerant nitrobacteria that enrichment obtains is back to incubator;
(2) cultivation-separation phase: after the Rich Internet Applications stage terminates, prepares initial ammonia nitrogen mass concentration 400mg/L, salinity massfraction in the growth nutrient solution of NaCl≤10% with water, flows into incubator by volume pump; The alkali lye of preparation massfraction 1% ~ 5%, flows into incubator by volume pump; After adding growth nutrient solution, carry out the pilot scale culture of nitrobacteria; Cultivate the mud mixed liquid obtained to be separated by precipitate and separate device, a bottom part collects the salt tolerant nitrobacteria of high density, and a part of salt tolerant nitrobacteria mud mixed liquid is back to incubator in addition, and supernatant liquor is discharged from precipitate and separate device top;
The condition of Rich Internet Applications stage and cultivation-separation phase is as follows: temperature is 10 DEG C ~ 35 DEG C, and pH is 6.0 ~ 10.0; DO is 1.5 ~ 7.5 mg/L.
2. the large-scaled culture method of the high density salt tolerant nitrobacteria of a kind of continous way according to claim 1, is characterized in that: composition and the consumption of described enrichment nutritive medium are: the ammonium salt of 0.5 ~ 10g/L, the KH of 0.5 ~ 5.0 g/L
2pO
4or K
2hPO
4, the MgCl of 0.5 ~ 5.0 g/L
2, the Na of 0.5 ~ 5.0 g/L
2cO
3, the NaCl of massfraction≤10%, liquid microelement;
The composition of described growth nutrient solution and consumption are: 0.5 ~ 10g/Lr ammonium salt, the KH of 0.5 ~ 10g/L
2pO
4or K
2hPO
4, the MgCl of 0.5 ~ 10g/L
2, the Na of 0.5 ~ 10g/L
2cO
3, the glucose of 0.5 ~ 10g/L or methyl alcohol, the NaCl of massfraction≤10%, liquid microelement;
Composition and the consumption thereof of described liquid microelement are as follows: the CoCl of 50 ~ 150mg/L
26H
2the CaCl of O, 50 ~ 150mg/L
2, the CuSO of 10 ~ 50mg/L
45H
2the ZnSO of O, 50 ~ 150mg/L
47H
2the FeSO of O, 50 ~ 150mg/L
47H
2the MnSO of O, 50 ~ 150mg/L
42H
2o.
3. the large-scaled culture method of the high density salt tolerant nitrobacteria of a kind of continous way according to claim 2, is characterized in that: described ammonium salt is selected from one or more of ammonium sulfate, urea or ammonium chloride.
4. the large-scaled culture method of the high density salt tolerant nitrobacteria of a kind of continous way according to claim 1, is characterized in that: according to step (1) described enriching method, every 1 ~ 2d is 1 enrichment cycle, and the enrichment cycle is 3 ~ 5; According to the cultural method described in step (2), every 2d ~ 3d is 1 culture cycle, and culture cycle is 5 ~ 8.
5. according to the large-scaled culture method of the high density salt tolerant nitrobacteria of a kind of continous way of claim 1-4 described in any one, it is characterized in that: the Rich Internet Applications stage adopts the method progressively improving ammonia nitrogen concentration, wherein ammonia nitrogen starting point concentration is 50 ~ 150mg/L, and ultimate density is 500 ~ 1000mg/L; Concrete grammar is as follows: after adding enrichment nutritive medium at every turn, when ammonia nitrogen concentration is down to below 5mg/L, improves the ammonia nitrogen concentration in enrichment nutritive medium, increase rate 20 ~ 100 mg/L; The dosage of suitable raising alkali lye, to guarantee the pH value environment required for nitrobacteria growth.
6., according to the large-scaled culture method of the high density salt tolerant nitrobacteria of a kind of continous way of claim 1-4 described in any one, it is characterized in that: cultivation-separation phase adopts stable ammonia nitrogen concentration, carry out concentration range correction simultaneously; Wherein stable ammonia nitrogen concentration is 400mg/L, and concentration correction scope is 300 ~ 500mg/L; Concrete grammar is as follows: add ammonia nitrogen concentration in grown cultures liquid is 400mg/L at every turn, as ammonia nitrogen concentration≤5mg/L, suitably improves ammonia nitrogen concentration and corrects, increase rate 10 ~ 100mg/L; As ammonia nitrogen concentration >=15 mg/L, suitably reduce ammonia nitrogen concentration and correct, reduce amplitude 10 ~ 100mg/L.
7., according to the large-scaled culture method of the high density salt tolerant nitrobacteria of a kind of continous way of claim 1-4 described in any one, it is characterized in that: the salt tolerant nitrobacteria mud mixed liquid volume fraction ratio that cultivation-separation phase is back to incubator bottom precipitate and separate device controls 10% ~ 40%.
8., according to the large-scaled culture method of the high density salt tolerant nitrobacteria of a kind of continous way of claim 1-4 described in any one, it is characterized in that: the ratio of the backflow flow velocity of cultivation-separation phase from precipitate and separate device to incubator controls at 0.1 ~ 1:1.
9. according to the large-scaled culture method of the high density salt tolerant nitrobacteria of a kind of continous way of claim 1-4 described in any one, it is characterized in that: Rich Internet Applications stage and cultivation-separation phase alkali lye used are NaOH, Na
2cO
3, NaHCO
3and CaCO
3in solution one or more.
10., according to the large-scaled culture method of the high density salt tolerant nitrobacteria of a kind of continous way of claim 1-4 described in any one, it is characterized in that: within Rich Internet Applications stage and cultivation-separation phase incubator, add appropriate active carbon powder.
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| CN106635947B (en) * | 2017-01-23 | 2021-03-23 | 中蓝连海设计研究院有限公司 | Asynchronous domestication culture method for salt-tolerant denitrification microbial inoculum |
| CN106635923B (en) * | 2017-01-23 | 2021-06-29 | 中蓝连海设计研究院有限公司 | Preparation method of high-density salt-tolerant denitrifying bacteria agent suitable for wastewater treatment |
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| CN114105294B (en) * | 2021-10-28 | 2023-09-22 | 苏州水星环保工业系统有限公司 | Construction and use method of high-salt high-ammonia-nitrogen organic wastewater biological treatment system |
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