A method for treating high-salinity low-temperature wastewater with nitrifying bacteria and denitrifying bacteria

Abstract

The application discloses a method for treating high-salt and low-temperature wastewater by using nitrifying bacteria and denitrifying bacteria, and adopts Paracoccus denitrificans TD-20229 screened from ammonia-nitrogen contaminated water bodies to simultaneously and cooperatively ferment wastewater in a high-salt and low-temperature environment together with Alcaligenes faecalis FC-01052. The production strain involved in the application can not only grow in a high-salt and low-temperature wastewater environment, but also has better denitrogenation effect. Compared with traditional physical and chemical denitrogenation methods, the production strain has the problems of low cost and secondary pollution reduction. The application obviously reduces wastewater pollution from the biological denitrogenation technology by using heterotrophic nitrifying bacteria and aerobic denitrifying bacteria, solves the problem of pesticide wastewater treatment in a low-temperature and high-salt environment, and has a wide market prospect.

Description

TECHNICAL FIELD

[0001] The present application relates to the field of biotechnology, in particular to a method for treating high-salt and low-temperature wastewater by using nitrifying bacteria and denitrifying bacteria. BACKGROUND

[0002] In recent years, human living standards have rapidly improved, the economy has developed rapidly, human production activities are frequent, and water environments have been affected by human activities. River pollution is serious, and the concentration of ammonia nitrogen in water bodies is also rapidly rising. Water bodies are in a state of eutrophication, and rivers are rich in excessive nitrogen content, causing serious water pollution of water resources in China and great harm to the safety of the ecological system. Therefore, improving water resources and finding an efficient denitrification method are key to the safety of water quality treatment. Controlling the nitrogen content in water bodies is the key to water pollution control, and the key to wastewater deep treatment and recovery. Traditional denitrification methods have the disadvantages of long process, high cost, high energy consumption, and the need to add alkaline substances for neutralization. In recent years, research on denitrifying microorganisms has become more in-depth, and heterotrophic nitrification-aerobic denitrification simultaneous reaction as a new type of biological denitrification technology has also been gradually applied to the treatment of wastewater pollution. It is also an economic, efficient, and environmentally friendly denitrification process.

[0003] Heterotrophic nitrification-aerobic denitrification bacteria break through the need for oxygen in the traditional biological nitrification and denitrification process, strict control and limitation of oxygen, and can complete the nitrification and denitrification process in the same reactor, can remove COD and reduce the content of ammonia nitrogen, control the nitrogen content, do not need to adjust the acid-base, and at the same time can repair the water environment through its own growth metabolism and adsorption, mineralization of pollutants. Under aerobic conditions, nitrifying bacteria will degrade and convert NH4 + -N or organic nitrogen into nitrite nitrogen and nitrate nitrogen, and then complete the denitrification process through denitrifying bacteria. Heterotrophic nitrifying bacteria have the advantages of strong adaptability, can grow with various organic matter as carbon source and other characteristics and are widely used by researchers. Aerobic denitrifying bacteria are a type of denitrifying bacteria that use aerobic denitrification enzymes to perform denitrification under aerobic conditions. Adding cultured aerobic denitrifying bacteria to wastewater treatment facilities can improve the denitrification effect of wastewater. Aerobic denitrifying bacteria can effectively and quickly remove nitrate in polluted water bodies. However, microbial denitrification is a series of denitrification reactions catalyzed by enzymes, which is affected by many factors, including wastewater temperature, dissolved oxygen, pH value, external carbon source, trace elements, etc. Among them, dissolved oxygen, pH value, inoculum size, and carbon source can be adjusted by process to improve denitrification effect. Suitable denitrification conditions are the key conditions for promoting the growth and metabolism of nitrifying bacteria and denitrifying bacteria and improving denitrification efficiency. Currently, existing technologies disclose that related nitrifying bacteria and denitrifying bacteria have the problems of poor denitrification effect, inability to grow and reproduce rapidly in low-temperature environments and alkaline environments, etc., thus reducing the removal rate of ammonia nitrogen, NO2 --N, NO 3- The removal effect of -N. The nitrobacteria used in the prior art include Pseudomonas, Acinetobacter, and Nitrosomonas, and the denitrifying bacteria include Bacillus, Paracoccus, and Pseudomonas. The existing strains have certain removal effect on ammonium nitrogen, nitrate nitrogen, and nitrite nitrogen, but have poor growth and denitrification effect in a low-temperature high-salt environment. SUMMARY

[0004] The application discloses a method for treating high-salt low-temperature wastewater by using nitrobacteria and denitrifying bacteria. - -N, NO 3- -N, and the denitrification effect is more obvious, thereby further achieving the purpose of treating wastewater. Compared with the traditional biological denitrification technology, the method has obvious denitrification effect, can reduce the content of ammonia nitrogen, NO2 - -N, NO 3- -N, and can grow well in a low-temperature high-salt environment. The method has the advantages of low price, simplified process, and solving the problem of wastewater treatment in a low-temperature high-salt environment to a certain extent.

[0005] To achieve the above object, the technical scheme adopted by the application is as follows:

[0006] A method for treating high-salt low-temperature wastewater by using nitrobacteria and denitrifying bacteria, comprising the following steps:

[0007] S1, activating the nitrobacteria, inoculating seed liquid, and culturing for 16-36h, inoculating into the fermentation medium according to the inoculation amount of 1-10%, the fermentation temperature is 5-42 DEG C, and the fermentation time is 48h-72h;

[0008] S2, activating the denitrifying bacteria, inoculating seed liquid, and culturing for 16-36h, inoculating into the fermentation medium according to the inoculation amount of 1-10%, the fermentation temperature is 5-42 DEG C, and the fermentation time is 48h-60h;

[0009] S3, adding the fermentation liquid of the two to the wastewater to be treated, and inoculating the nitrobacteria and the denitrifying bacteria according to the volume ratio of 1-10:1-10, first adding the nitrobacteria to treat the wastewater for 10-18h, and then adding the denitrifying bacteria to treat for 30-48h.

[0010] The nitrobacteria are Alcaligenes FC-01052, and the denitrifying bacteria are Paracoccus TD-20229.

[0011] The Alcaligenes sp. FC-01052 is preserved in the China General Microbiological Culture Collection Center, and the preservation number is CGMCC No. 24294; the Paracoccus denitrificans TD-20229 has been preserved in the China General Microbiological Culture Collection Center on September 15, 2022, and the address is No. 3, Beichen West Road, Haidian District, Beijing, and the preservation number is CGMCC No. 25718.

[0012] The seed liquid component is: glucose 5 g / L, peptone 10 g / L, yeast powder 5 g / L, sodium chloride 10 g / L, pH 7.

[0013] The nitrifying bacteria culture medium component is: carbon source 5-30 g / L, nitrogen source 5-30 g / L, inorganic salt 1-10 g / L, metal ion 0-2 g / L, trace element 0.1-1 g / L, the rest is water, pH 4-9.0.

[0014] The denitrifying bacteria culture medium component is: carbon source 5-30 g / L, nitrogen source 5-30 g / L, inorganic salt 1-10 g / L, metal ion 0-2 g / L, trace element 0.1-1 g / L, the rest is water, pH 4-9.0.

[0015] The carbon source is at least one of glucose, trisodium citrate, sodium pyruvate, sodium acetate, molasses, etc.

[0016] The nitrogen source is at least one of ammonium chloride, ammonium nitrate, sodium nitrite, sodium nitrate, potassium nitrate, ammonium sulfate, urea, beef extract, ammonia water, corn syrup, fish meal, etc.

[0017] The inorganic salt is any one or a combination of several of magnesium salt, nitrate, potassium salt, calcium salt, phosphate or hydrochloride.

[0018] The trace element is a mixture of ferrous sulfate 0.01-3 g / L, copper sulfate 0.001-1 g / L, zinc sulfate 0.01-0.5 g / L, calcium chloride 0.1-2 g / L, manganese sulfate 0.01-1 g / L, and cobalt chloride 0.001-0.1 g / L.

[0019] Further preferably, the nitrifying bacteria are activated, inoculated with seed liquid, cultured for 24 h, inoculated into the fermentation medium at a inoculation amount of 3%, and fermented at 37°C for 48 h;

[0020] The denitrifying bacteria are activated, inoculated with seed liquid, cultured for 24 h, inoculated into the fermentation medium at a inoculation amount of 3%, and fermented at 37°C for 50 h;

[0021] The two fermentation liquids are added into the wastewater to be treated, and the nitrifying bacteria and the denitrifying bacteria are added in a volume ratio of 1:1, the nitrifying bacteria are added first to treat the wastewater for 10-18 hours, and then the denitrifying bacteria are added to treat the wastewater for 30-48 hours.

[0022] More specifically, the Alcaligenes FC-01052 and the Paracoccus denitrificans TD-20229 are activated respectively, inoculated into seed liquid (glucose 5g / L, peptone 10g / L, yeast powder 5g / L, sodium chloride 10g / L, pH 7), and cultured for 24 hours, and then inoculated into the same fermentation medium in a 3% inoculation amount, and fermented at 15-45℃ for 48-84 hours, and the aeration ratio is 1.0-2.5 v / vm and the stirring speed is 150-200 rpm during the fermentation, wherein the fermentation medium is composed of the following components: trisodium citrate 20g / L, ammonium nitrate 4.5g / L, potassium nitrate 3g / L, NaNO2 0.3g / L, urea 1g / L, Na2HPO4·3H2O 2g / L, NaH2PO4 2g / L, MgSO4·7H2O 0.1g / L, ferrous sulfate 0.01g / L, copper sulfate 0.002g / L, zinc sulfate 0.01g / L, calcium chloride 0.5g / L, manganese sulfate 0.03g / L, cobalt chloride 0.002g / L, and the rest is water, and the pH is 7.5. When the fermentation is completed, the fermentation liquid is added into the wastewater to be treated in a volume ratio of 1:1, and the wastewater is treated, wherein the treatment mode is that the Alcaligenes FC-01052 is added first to treat the wastewater for 15 hours, and then the Paracoccus denitrificans TD-20229 is added to treat the wastewater for 40 hours.

[0023] The added amount of the fermentation liquid into the wastewater to be treated is 2%-10%.

[0024] The Alcaligenes FC-01052 and the Paracoccus denitrificans TD-20229 can grow well in a low-temperature and high-salt environment and have obvious denitrification ability.

[0025] The Alcaligenes FC-01052 has obvious degradation effect on ammonia nitrogen in wastewater in a low-temperature and high-salt wastewater environment, and the degradation rate is more than 90%. The ammonia nitrogen degradation function of the strain is reported for the first time.

[0026] The Paracoccus denitrificans TD-20229 has obvious degradation effect on ammonia nitrogen, NO2 - -N, and NO3 - -N in wastewater in a low-temperature and high-salt wastewater environment, and the degradation rates are more than 90%, more than 99%, and more than 99%, respectively.

[0027] Paracoccus denitrificans TD-20229 is screened by the following method: five strains of Paracoccus denitrificans are isolated from a sewage plant, basic culture is carried out by using liquid LB culture medium, after 1 day of culture at 35 DEG C, the biomass is determined at the same time, a strain with the most vigorous activity is obtained, named as TD-1, as a dominant strain.

[0028] TD-1 is inoculated in liquid LB seed culture medium for culture, and the bacterial liquid of the cultured Paracoccus denitrificans is diluted to a cell number of 10 6 CFU / mL by 10-fold dilution method, 1 mL of the bacterial liquid is uniformly coated on a sterile empty flat plate, after air drying, N + ion beam implantation, N + The ion beam implantation dose is (90, 135, 180, 225, 270) x 2.6 x 10 13 N + / cm 2 , N + The ion beam implantation energy is 20 keV. After irradiation, the cells are washed with 1 mL of sterile water, diluted by 10-fold dilution method, and coated into flat plate culture medium, and cultured at 37 DEG C for 1 day, after picking single colonies, inoculated into LB seed liquid, cultured at 37 DEG C and 150 rpm for 12 h, and then cultured in nitration medium (NH4 + -N medium, NO3 - -N medium and NO2 - -N medium) with a 5% inoculation amount, respectively, to screen out a strain with better colony growth and degradation effect, named as Paracoccus denitrificans TD-20229. The strain grows rapidly under aerobic culture, grows slowly under anoxic condition, the suitable growth temperature is 25-30 DEG C, and the growth pH is 6.5-9; the spherical cells (diameter 0.5-0.9 um) or short rods (length 0.9-1.2 um) are single, paired or stacked, form poly-beta-hydroxybutyrate particles, gram-negative, non-motile, aerobic and respiratory metabolism.

[0029] In the present application, after the fermentation of Alcaligenes FC-01052 and Paracoccus denitrificans TD-20229 is completed, the fermentation liquid is added into the wastewater to be treated according to a volume ratio of 1:1, and the wastewater is treated, wherein the treatment mode is that the wastewater is first treated by adding Alcaligenes FC-01052, and then treated by adding Paracoccus denitrificans TD-20229.

[0030] The high-salt (1-10%) and low-temperature (5 DEG C-20 DEG C) sewage in the present application is pesticide and chemical wastewater.

[0031] The application mainly relates to the culture, fermentation process optimization of Alcaligenes and Paracoccus denitrificans and the application in the treatment of low-temperature and high-salt sewage, avoids secondary pollution, has a denitrification effect and further achieves the purpose of treating sewage. Compared with the prior art, the method of the application has the following advantages: after the fermentation of Alcaligenes FC-01052 and Paracoccus denitrificans TD-20229 is completed, the fermentation liquor is added into the wastewater to be treated according to a volume ratio of 1:1, the wastewater is treated, the denitrification effect is obvious, the ammonia nitrogen, NO2 - -N, NO3 - -N, and can grow well in a low-temperature and high-salt environment. The production strain for removing ammonia nitrogen solves the problem of wastewater treatment and further provides a theoretical basis and technical guidance for solving related problems.

[0032] Beneficial effects: compared with the prior art, the application has the following advantages: the application uses Alcaligenes FC-01052 and Paracoccus denitrificans TD-20229, combines the characteristics that the Alcaligenes can efficiently degrade ammonia nitrogen in a low-temperature and high-salt environment, and cooperates the Paracoccus denitrificans to synergistically and efficiently degrade ammonia nitrogen, NO 2- -N, NO 3- -N in a low-temperature and high-salt pesticide wastewater environment, and the denitrification effect is more obvious.

[0033] 1. Compared with other strains, the Alcaligenes and Paracoccus denitrificans have high metabolic activity, can grow well in a low-temperature and high-salt environment, have good denitrification effect and good wastewater treatment effect.

[0034] 2. Compared with the traditional nitrification-denitrification denitrification process, the process mixes the Alcaligenes and Paracoccus denitrificans to reduce the nitrogen content in the wastewater, does not need to neutralize acid and alkali, simplifies the process operation and saves energy consumption and the like. DETAILED DESCRIPTION

[0035] The application can be better understood through the following examples. Then, those skilled in the art can easily understand that the specific material ratio, process conditions and results described in the examples are only used to illustrate the application and should not and will not limit the application described in detail in the claims.

[0036] Example 1: Influence of temperature on strain growth and denitrification effect

[0037] Alcaligenes faecalis FC-01052 and Paracoccus denitrificans TD-20229 were activated respectively, inoculated into seed liquid (10 g / L of proteose peptone, 5 g / L of yeast powder, 10 g / L of sodium chloride, 5 g / L of glucose, pH 7), and cultured for 20 h. Then, the two strains were respectively inoculated into fermentation medium at a ratio of 3%, and fermented for 48 h at 5, 10, 15, 20, 25, 30, 37 and 42 ℃, with aeration ratio of 1.5 v / vm and stirring at 180 rpm. The fermentation medium for the two strains was composed of 20 g / L of trisodium citrate, 4.5 g / L of ammonium nitrate, 3 g / L of potassium nitrate, 0.3 g / L of NaNO2, 1 g / L of urea, 2 g / L of Na2HPO4·3H2O, 2 g / L of NaH2PO4, 0.1 g / L of MgSO4·7H2O, 0.01 g / L of ferrous sulfate, 0.002 g / L of copper sulfate, 0.01 g / L of zinc sulfate, 0.5 g / L of calcium chloride, 0.03 g / L of manganese sulfate, 0.002 g / L of cobalt chloride, and water, with pH 7.5. The effects of different temperatures on the growth of the strains and the denitrification effect were investigated.

[0038] At the end of fermentation, the fermentation liquid was respectively inoculated into 2% nitrification medium (NH4 + -N medium, NO3 - -N medium and NO2 - -N medium), and aerobically fermented for 48 h at 35 ℃. The denitrification test and growth condition were respectively carried out in the three different media.

[0039] The NH4 + -N simulated wastewater medium: 5 g / L of sodium acetate, 0.8 g / L of (NH4)2SO4, 0.5 g / L of NaCL, 1 g / L of disodium hydrogen phosphate, 1 g / L of sodium dihydrogen phosphate, 0.02 g / L of FeSO4·7H2O, 0.05 g / L of MgSO4·7H2O, 0.01 g / L of zinc sulfate, and 0.01 g / L of manganese chloride, with pH 7.2.

[0040] The NaNO3 simulated wastewater medium: 5 g / L of sodium acetate, 0.8 g / L of KNO3, 0.5 g / L of NaCL, 1 g / L of disodium hydrogen phosphate, 1 g / L of sodium dihydrogen phosphate, 0.02 g / L of FeSO4·7H2O, 0.05 g / L of MgSO4·7H2O, 0.01 g / L of zinc sulfate, and 0.01 g / L of manganese chloride, with pH 7.2.

[0041] NaNO2 simulated wastewater medium: NaNO20.8 g / L, sodium acetate 5 g / L, NaCL 0.5 g / L, Na2HPO41 g / L, NaH2PO11 g / L, FeSO4·7H2O 0.02 g / L, MgSO4·7H2O 0.05 g / L, ZnSO40.01 g / L, MnCL0.01 g / L, pH 7.2.

[0042] Temperature has a great influence on microbial nitrification and denitrification. As shown in Tables 1 and 2, temperature has a great influence on the nitrification and denitrification activities of Alcaligenes sp. FC-01052 and Paracoccus denitrificans TD-20229. When the temperature is 5-15℃, the NO3 - -N, NO2 - The removal rate of -N is more than 40%, and the removal rate of ammonia nitrogen gradually increases with the increase of temperature. When the temperature is 20-30℃, the removal rate of ammonia nitrogen is more than 80%; when the temperature is 35℃, the removal rate of ammonia nitrogen is more than 92%; when the temperature is 42℃, the removal rate of total nitrogen decreases, and the nitrification and denitrification abilities decrease obviously, indicating that the growth and denitrification effect of the two strains are better in low-temperature environment. As shown in Tables 1 and 2, under the same temperature condition, the degradation ability of ammonia nitrogen of Alcaligenes sp. FC-01052 is much higher than that of Paracoccus denitrificans TD-20229, and the ability of Paracoccus denitrificans TD-20229 to remove NO3 - -N, NO2 - The ability of Paracoccus denitrificans TD-20229 to remove NO3

[0043] Table 1 Influence of temperature on the growth and denitrification effect of Alcaligenes sp. FC-01052

[0044]

[0045]

[0046] Table 2 Influence of temperature on the growth and denitrification effect of Paracoccus denitrificans TD-20229

[0047]

[0048] Example 2 Influence of high salt on the growth and denitrification effect of strains

[0049] Alcaligenes FC-01052 and Paracoccus TD-20229 were activated respectively, inoculated into seed liquid (10 g / L of proteose peptone, 5 g / L of yeast powder, 10 g / L of sodium chloride, 5 g / L of glucose, pH 7), and cultured for 20 h. Then, the two strains were inoculated into fermentation medium respectively at a ratio of 3%, and fermented for 48 h under the conditions of 1%, 2%, 4%, 6%, 8%, and 10% of sodium chloride concentration, 1.5 v / vm of aeration ratio, and 180 rpm of stirring. The fermentation medium for the two strains was composed of 20 g / L of trisodium citrate, 4.5 g / L of ammonium nitrate, 3 g / L of potassium nitrate, 0.3 g / L of NaNO2, 1 g / L of urea, 2 g / L of Na2HPO4·3H2O, 2 g / L of NaH2PO4, 0.1 g / L of MgSO4·7H2O, 0.01 g / L of ferrous sulfate, 0.002 g / L of copper sulfate, 0.01 g / L of zinc sulfate, 0.5 g / L of calcium chloride, 0.03 g / L of manganese sulfate, and 0.002 g / L of cobalt chloride, and the rest was water, with pH being 7.5. The effects of different salinity on the growth of the strains and the denitrification effect were investigated.

[0050] After the fermentation, the fermentation liquid was inoculated into 2% nitrification medium (NH4 + -N medium, NO3 - -N medium, and NO2 - -N medium), and aerobically fermented for 48 h at 35°C. The denitrification test and the growth condition were carried out in the three different media respectively.

[0051] The NH4 + -N simulated wastewater medium: 5 g / L of sodium acetate, 0.8 g / L of (NH4)2SO4, 0.5 g / L of NaCL, 1 g / L of Na2HPO4, 1 g / L of NaH2PO4, 0.02 g / L of FeSO4·7H2O, 0.05 g / L of MgSO4·7H2O, 0.01 g / L of ZnSO4, and 0.01 g / L of MnCL2, with pH being 7.2.

[0052] The NaNO3 simulated wastewater medium: 5 g / L of sodium acetate, 0.8 g / L of KNO3, 0.5 g / L of NaCL, 1 g / L of Na2HPO4, 1 g / L of NaH2PO4, 0.02 g / L of FeSO4·7H2O, 0.05 g / L of MgSO4·7H2O, 0.01 g / L of ZnSO4, and 0.01 g / L of MnCL2, with pH being 7.2.

[0053] NaNO2 simulated wastewater medium: NaNO20.8 g / L, sodium acetate 5 g / L, NaCL 0.5 g / L, Na2HPO41 g / L, NaH2PO11 g / L, FeSO4·7H2O 0.02 g / L, MgSO4·7H2O 0.05 g / L, ZnSO40.01 g / L, MnCL2 0.01 g / L, pH 7.2.

[0054] The salinity has a great influence on the microbial nitrification and denitrification. The following tables show that the salinity has a great influence on the nitrification and denitrification of Alcaligenes FC-01052 and Paracoccus denitrificans TD-20229. The tables 3 and 4 also show that the ammonia nitrogen degradation ability of Alcaligenes FC-01052 is much higher than that of Paracoccus denitrificans TD-20229 under the same salinity. When the salinity is 1-10%, the removal rate of NO3 - -N, NO2 - The removal rate of -N is more than 40%, and the removal rate of ammonia nitrogen is gradually increased with the increase of salinity. When the salinity is 1-6%, the removal rate of ammonia nitrogen is more than 80%. When the salinity is 1-4%, the removal rate of ammonia nitrogen is more than 90%. When the salinity is higher than 4%, the removal rate of NO3 - -N, NO2 - The removal rate of -N is decreased, and the nitrification and denitrification ability is obviously decreased, which shows that the growth and denitrification effect of the two strains are better in the low temperature environment. The tables 3 and 4 also show that the ammonia nitrogen degradation ability of Alcaligenes FC-01052 is much higher than that of Paracoccus denitrificans TD-20229 under the same salinity. The NO3 - -N, NO2 - The NO3

[0055] Table 3 Influence of salinity on the growth and denitrification effect of Alcaligenes FC-01052

[0056]

[0057]

[0058] Table 4 Influence of salinity on the growth and denitrification effect of Paracoccus denitrificans TD-20229

[0059]

[0060] Effect of initial pH on strain growth and denitrification

[0061] Alcaligenes FC-01052 and Paracoccus TD-20229 were activated respectively and inoculated into seed liquid (10 g / L proteose peptone, 5 g / L yeast powder, 10 g / L sodium chloride, 5 g / L glucose, pH 7) and cultured for 20 h. Then, according to the inoculation amount of 3%, the strains were transferred into fermentation medium and fermented for 48 h at 4, 4.5, 5, 5.5, 6, 6.5, 7, 7.5, 8, 9 ℃ respectively. During the fermentation stage, the aeration ratio was 1.5 v / vm and the stirring was 180 rpm. The fermentation medium for Alcaligenes and Paracoccus was as follows: 20 g / L trisodium citrate, 4.5 g / L ammonium nitrate, 3 g / L potassium nitrate, 0.3 g / L NaNO2, 1 g / L urea, 2 g / L Na2HPO4·3H2O, 2 g / L NaH2PO4, 0.1 g / L MgSO4·7H2O, 0.01 g / L ferrous sulfate, 0.002 g / L copper sulfate, 0.01 g / L zinc sulfate, 0.5 g / L calcium chloride, 0.03 g / L manganese sulfate, 0.002 g / L cobalt chloride, and the rest was water, pH 7.5. The effect of different initial pH on strain growth and denitrification was investigated. Tables 5 and 6 show that under the same pH condition, the ammonia nitrogen degradation ability of Alcaligenes FC-01052 is much higher than that of Paracoccus TD-20229, and the NO 3- -N, NO 2- -N, and the ability of Paracoccus TD-20229 to remove NO

[0062] At the end of fermentation, the fermented fermentation liquid was respectively put into 2% nitrification medium (NH + -N medium, NO3 - -N medium and NO2 - -N medium), 35 ℃ aerobic fermentation for 48 h, and denitrification test and growth were carried out in three different media respectively.

[0063] Among them, NH + -N simulated wastewater medium: sodium acetate 5 g / L, (NH4)2SO4 0.8 g / L, NaCL 0.5 g / L, disodium hydrogen phosphate 1 g / L, sodium dihydrogen phosphate 1 g / L, FeSO4·7H2O 0.02 g / L, MgSO4·7H2O 0.05 g / L, zinc sulfate 0.01 g / L, manganese chloride 0.01 g / L, pH 7.2.

[0064] NaNO3 simulated wastewater medium: NaNO3 0.8 g / L, NaAc 5 g / L, NaCl 0.5 g / L, Na2HPO4 1 g / L, NaH2PO4 1 g / L, FeSO4.7H2O 0.02 g / L, MgSO4.7H2O 0.05 g / L, ZnSO4 0.01 g / L, MnCl2 0.01 g / L, pH 7.2.

[0065] NaNO2 simulated wastewater medium: NaNO2 0.8 g / L, NaAc 5 g / L, NaCl 0.5 g / L, Na2HPO4 1 g / L, NaH2PO4 1 g / L, FeSO4.7H2O 0.02 g / L, MgSO4.7H2O 0.05 g / L, ZnSO4 0.01 g / L, MnCl2 0.01 g / L, pH 7.2.

[0066] The initial pH has a great influence on the microbial nitrification and denitrification. As shown in Tables 5 and 6, the temperature has a great influence on the nitrification and denitrification activities of Alcaligenes sp. FC-01052 and Paracoccus denitrificans TD-20229. When the initial pH is 4-9, the NO3 - -N, NO2 - The removal rate of -N is more than 40%, and the removal rate of ammonia nitrogen gradually increases with the increase of pH. When the initial pH is 6-8, the removal rate of ammonia nitrogen is more than 80%; when the initial pH is 4-5, the removal rate of ammonia nitrogen is more than 60%; when the pH is higher than 8, the removal rate of total nitrogen decreases, and the nitrification and denitrification abilities obviously decrease, indicating that the growth and denitrification effect of the two strains are better in low-temperature environment. As shown in Tables 5 and 6, under the same pH condition, the degradation ability of ammonia nitrogen of Alcaligenes sp. FC-01052 is much higher than that of Paracoccus denitrificans TD-20229, and the denitrification ability of Paracoccus denitrificans TD-20229 is obviously higher than that of Alcaligenes sp. FC-01052, especially when the initial pH is 6.5-8, the degradation ability of ammonia nitrogen of Alcaligenes sp. FC-01052 is more than 90%, and the degradation rate is 90%-99.8%; when the initial pH is 4-6.5, the degradation ability of ammonia nitrogen is more than 60%, and the degradation rate is 60.4%-89.9%. - -N, NO2 - The removal rate of -N is more than 40%, and the removal rate of ammonia nitrogen gradually increases with the increase of pH. When the initial pH is 6-8, the removal rate of ammonia nitrogen is more than 80%; when the initial pH is 4-5, the removal rate of ammonia nitrogen is more than 60%; when the pH is higher than 8, the removal rate of total nitrogen decreases, and the nitrification and denitrification abilities obviously decrease, indicating that the growth and denitrification effect of the two strains are better in low-temperature environment. As shown in Tables 5 and 6, under the same pH condition, the degradation ability of ammonia nitrogen of Alcaligenes sp. FC-01052 is much higher than that of Paracoccus denitrificans TD-20229, and the denitrification ability of Paracoccus denitrificans TD-20229 is obviously higher than that of Alcaligenes sp. FC-01052, especially when the initial pH is 6.5-8, the degradation ability of ammonia nitrogen of Alcaligenes sp. FC-01052 is more than 90%, and the degradation rate is 90%-99.8%; when the initial pH is 4-6.5, the degradation ability of ammonia nitrogen is more than 60%, and the degradation rate is 60.4%-89.9%.

[0067] Table 5 Influence of initial pH on the growth and denitrification effect of Alcaligenes sp. FC-01052

[0068]

[0069]

[0070] Table 6 Influence of initial pH on the growth and denitrification effect of Paracoccus denitrificans TD-20229

[0071]

[0072] Example 4 Influence of carbon source on strain growth and denitrification effect

[0073] Alcaligenes FC-01052 and Paracoccus denitrificans TD-20229 were activated respectively and inoculated into seed liquid (10 g / L proteose peptone, 5 g / L yeast powder, 10 g / L sodium chloride, 5 g / L glucose, pH 7) and cultured for 20 h. Then, according to the inoculation amount of 3%, the strains were transferred into fermentation medium and fermented for 48 h under carbon source conditions (sodium citrate, sodium acetate, sodium pyruvate). During the fermentation stage, the aeration ratio was 1.5 v / vm and the stirring was 180 rpm. The fermentation medium for Alcaligenes and Paracoccus denitrificans was as follows: 20 g / L trisodium citrate, 4.5 g / L ammonium nitrate, 3 g / L potassium nitrate, 0.3 g / L NaNO2, 1 g / L urea, 2 g / L Na2HPO4·3H2O, 2 g / L NaH2PO4, 0.1 g / L MgSO4·7H2O, 0.01 g / L ferrous sulfate, 0.002 g / L copper sulfate, 0.01 g / L zinc sulfate, 0.5 g / L calcium chloride, 0.03 g / L manganese sulfate, 0.002 g / L cobalt chloride, and the rest was water, pH 7.5. The influence of different carbon sources on strain growth and denitrification effect was investigated.

[0074] It can be seen from Tables 7 and 8 that under the carbon source conditions, the ammonia nitrogen degradation ability of Alcaligenes FC-01052 was much higher than that of Paracoccus denitrificans TD-20229, while the NO 3- -N, NO 2- -N removal ability of Paracoccus denitrificans TD-20229 was obviously higher than that of Alcaligenes FC-01052.

[0075] At the end of fermentation, the fermentation liquid was respectively poured into 2% nitrification medium (NH4 + -N medium, NO3 - -N medium and NO2 - -N medium), and aerobic fermentation was carried out at 35°C for 48 h. Denitrification test and growth condition were carried out in the three different media respectively.

[0076] Among them, the NH4 + -N simulated wastewater medium: 5 g / L sodium acetate, 0.8 g / L (NH4)2SO4, 0.5 g / L NaCL, 1 g / L disodium hydrogen phosphate, 1 g / L sodium dihydrogen phosphate, 0.02 g / L FeSO4·7H2O, 0.05 g / L MgSO4·7H2O, 0.01 g / L zinc sulfate, 0.01 g / L manganese chloride, pH 7.2.

[0077] NaNO3 simulated wastewater medium: NaNO3 0.8 g / L, NaAc 5 g / L, NaCl 0.5 g / L, Na2HPO4 1 g / L, NaH2PO4 1 g / L, FeSO4.7H2O 0.02 g / L, MgSO4.7H2O 0.05 g / L, ZnSO4 0.01 g / L, MnCl2 0.01 g / L, pH 7.2.

[0078] NaNO2 simulated wastewater medium: NaNO2 0.8 g / L, NaAc 5 g / L, NaCl 0.5 g / L, Na2HPO4 1 g / L, NaH2PO4 1 g / L, FeSO4.7H2O 0.02 g / L, MgSO4.7H2O 0.05 g / L, ZnSO4 0.01 g / L, MnCl2 0.01 g / L, pH 7.2.

[0079] The results in Tables 7 and 8 show that temperature has a great effect on the nitrification and denitrification activities of Alcaligenes sp. FC-01052 and Paracoccus denitrificans TD-20229. Under different carbon sources, the removal rates of NO3 - -N, NO2 - -N were all above 60%. When the nitrogen source was trisodium citrate, the removal rate of NO3 - -N, NO2 - -N by Alcaligenes sp. FC-01052 was obviously higher than that by using NaAc or Na-pyruvate as the carbon source. - -N, NO2 - -N. The removal rate of ammonia nitrogen was still 99.2%-99.8%. This shows that the growth and denitrification effect of the two strains are better in low temperature environment. Table 7 and Table 8 also show that the ability of Alcaligenes sp. FC-01052 to degrade ammonia nitrogen is much higher than that of Paracoccus denitrificans TD-20229, and the ability of Paracoccus denitrificans TD-20229 to remove NO3 - -N, NO2 - -N is higher than that of Alcaligenes sp. FC-01052.

[0080] Table 7 Effect of carbon source on the growth and denitrification effect of Alcaligenes sp. FC-01052

[0081]

[0082] Table 8 Effect of carbon source on the growth and denitrification effect of Paracoccus denitrificans TD-20229

[0083]

[0084] Example 5 Effect of different treatment methods on denitrification

[0085] Alcaligenes faecalis FC-01052 and Paracoccus denitrificans TD-20229 were activated respectively, inoculated into seed liquid (10 g / L proteose peptone, 5 g / L yeast powder, 10 g / L sodium chloride, 5 g / L glucose, pH 7), and cultured for 20 h. Then, according to the inoculation amount of 3%, the two strains were transferred into fermentation medium, and fermented for 48 h under carbon source conditions (sodium citrate, sodium acetate, sodium pyruvate). During the fermentation stage, the aeration ratio was 1.5 v / vm, and the stirring was 180 rpm. The fermentation medium for the two strains was as follows: 20 g / L trisodium citrate, 4.5 g / L ammonium nitrate, 3 g / L potassium nitrate, 0.3 g / L NaNO2, 1 g / L urea, 2 g / L Na2HPO4·3H2O, 2 g / L NaH2PO4, 0.1 g / L MgSO4·7H2O, 0.01-3 g / L ferrous sulfate, 0.002 g / L copper sulfate, 0.01 g / L zinc sulfate, 0.5 g / L calcium chloride, 0.03 g / L manganese sulfate, 0.002 g / L cobalt chloride, and the rest was water, pH 7.5. At the end of fermentation, the fermentation liquid was inoculated into the following simulated wastewater medium according to different treatment methods for denitrification.

[0086] At the end of fermentation, the fermentation liquid was inoculated into 2% nitrification medium (NH4 + -N medium, NO3 - -N medium and NO2 - -N medium), and aerobically fermented for 48 h at 35°C. Denitrification test and growth were carried out in the three different media, respectively.

[0087] Among them, NH4 + -N simulated wastewater medium: 5 g / L sodium acetate, 0.8 g / L (NH4)2SO4, 0.5 g / L NaCL, 1 g / L disodium hydrogen phosphate, 1 g / L sodium dihydrogen phosphate, 0.02 g / L FeSO4·7H2O, 0.05 g / L MgSO4·7H2O, 0.01 g / L zinc sulfate, 0.01 g / L manganese chloride, pH 7.2.

[0088] NaNO3 simulated wastewater medium: 5 g / L sodium acetate, 0.8 g / L KNO3, 0.5 g / L NaCL, 1 g / L disodium hydrogen phosphate, 1 g / L sodium dihydrogen phosphate, 0.02 g / L FeSO4·7H2O, 0.05 g / L MgSO4·7H2O, 0.01 g / L zinc sulfate, 0.01 g / L manganese chloride, pH 7.2.

[0089] NaNO2 simulated wastewater medium: NaNO2 0.8 g / L, sodium acetate 5 g / L, NaCL 0.5 g / L, Na2HPO4 1 g / L, NaH2PO4 1 g / L, FeSO4·7H2O 0.02 g / L, MgSO4·7H2O 0.05 g / L, ZnSO4 0.01 g / L, MnCL2 0.01 g / L, pH 7.2.

[0090] Treatment method ①: the fermentation broth was added to the wastewater to be simulated according to (volume ratio 1:1), and the wastewater was treated, wherein the treatment method was to first add Alcaligenes FC-01052 to treat the wastewater for 15 h, and then add Paracoccus TD-20229 to treat for 40 h.

[0091] Treatment method ②: the fermentation broth was added to the wastewater to be simulated according to (volume ratio 1:1) at the same time, and the wastewater was treated.

[0092] Treatment method ③: the two fermentation broths were added to the wastewater to be treated, and the simulated wastewater was treated.

[0093] Treatment method ④: the fermentation broth was added to the wastewater to be simulated according to (volume ratio 1:1), and the wastewater was treated, wherein the treatment method was to first add Paracoccus TD-20229 to treat for 15 h, and then add Alcaligenes FC-01052 to treat for 40 h.

[0094] Different treatment methods have obvious different effects on the denitrification of the two strains in the low-temperature high-salt wastewater environment. When the fermentation broth is added to the wastewater to be treated according to (volume ratio 1:1), and the wastewater is treated, first Alcaligenes FC-01052 is added to treat the wastewater for 15 h, and then Paracoccus TD-20229 is added to treat for 40 h. The removal rates of NO3 - -N, NO2 - -N and NH4 + -N are 99.7%, 99.2% and 99.3% respectively, and the removal effect of treatment method ① is the best.

[0095] Table 9 Effect of different treatment methods on denitrification

[0096]

[0097] Example 6 Study of strains in low-temperature high-salt industrial wastewater

[0098] Alcaligenes FC-01052 and Paracoccus denitrificans TD-20229 are respectively activated, inoculated into seed liquid (10 g / L of proteose peptone, 5 g / L of yeast powder, 10 g / L of sodium chloride, 10 g / L of glucose, pH 7), and cultured for 20 h, then inoculated into fermentation medium at a ratio of 3%, and fermented for 48 h under carbon source conditions (sodium citrate, sodium acetate, sodium pyruvate), with aeration ratio of 1.5 v / vm and stirring at 180 rpm in the fermentation stage, wherein the fermentation medium of the Alcaligenes and the Paracoccus denitrificans is: 20 g / L of trisodium citrate, 4.5 g / L of ammonium nitrate, 3 g / L of potassium nitrate, 0.3 g / L of NaNO2, 1 g / L of urea, 2 g / L of Na2HPO4·3H2O, 2 g / L of NaH2PO4, 0.1 g / L of MgSO4·7H2O, 0.01 g / L of ferrous sulfate, 0.002 g / L of copper sulfate, 0.01 g / L of zinc sulfate, 0.5 g / L of calcium chloride, 0.03 g / L of manganese sulfate, 0.002 g / L of cobalt chloride, and the rest is water, pH 7.5.

[0099] At the end of fermentation, the fermented fermentation liquid is inoculated into the fermentation medium at a ratio of 2%, and aerobically fermented at 35℃ for 48 h, and the fermentation liquid is added to the pesticide chemical wastewater at a ratio of 1:1 (volume ratio) for wastewater treatment, wherein the treatment method is that the Alcaligenes FC-01052 is added to the wastewater for 15 h, and then the Paracoccus denitrificans TD-20229 is added for 40 h.

[0100] The growth and denitrification effect of the strains in the low-temperature high-salt industrial wastewater are investigated under the conditions of temperature of 15℃, salinity of 4%, ammonia nitrogen concentration of 400 mg / L, NO3 - -N concentration of 300 mg / L, and NO2 - -N concentration of 300 mg / L. As shown in Table 10, the denitrification effect of the Alcaligenes FC-01052 and the Paracoccus denitrificans TD-20229 in the low-temperature high-salt wastewater is excellent, the removal rates of NO3 - -N, NO2 - -N, and NH4 + -N are 89.7%, 88.4%, and 90.6%, respectively.

[0101] Table 10 Influence of growth and denitrification effect of strains in low-temperature high-salt industrial wastewater

[0102]

[0103] The detection method of the process in the application: the concentration of NO3 - -N is determined by ultraviolet spectrophotometry, the concentration of NO2 - -N is determined by N-(1-naphthyl)-ethylenediamine spectrophotometry, and the concentration of NH4 +The -N concentration is determined by the Nessler's reagent spectrophotometry.

[0104] The above describes the technical solutions provided by the embodiments of the present application in detail. The principles and implementation manners of the embodiments of the present application are described by using specific examples. The above descriptions of the embodiments are only used to help understand the principles of the embodiments of the present application. For the general skilled in the art, the specific implementation manners and application scopes of the embodiments of the present application will be changed. In conclusion, the content of the present description should not be understood as a limitation of the present application.

Claims

1. A method for treating high-salinity low-temperature wastewater with nitrifying bacteria and denitrifying bacteria, characterized in that, It comprises the following steps: S1, activating nitrifying bacteria, inoculating seed liquid, culturing for 16-36 h, inoculating into fermentation medium according to inoculation amount of 1-10%, fermentation temperature of 5-42 DEG C, and fermentation time of 48 h-72 h; S2, activating denitrifying bacteria, inoculating seed liquid, culturing for 16-36 h, inoculating into fermentation medium according to inoculation amount of 1-10%, fermentation temperature of 5-42 DEG C, and fermentation time of 48 h-60 h; S3, adding the fermentation liquid of the two to the wastewater to be treated, adding nitrifying bacteria first according to volume ratio of nitrifying bacteria to denitrifying bacteria of 1-10:1-10, treating the wastewater for 10-18 h, then adding denitrifying bacteria and treating for 30-48 h; The nitrifying bacteria are Alcaligenes FC-01052, the preservation number of which is CGMCC No.24294; the denitrifying bacteria are Paracoccus denitrificans TD-20229, the preservation number of which is CGMCC No.25718. The high-salt low-temperature wastewater is pesticide chemical wastewater with salt concentration of 1-10% and temperature of 5 DEG C-20 DEG C.

2. The method for treating high-salinity low-temperature wastewater with nitrifying bacteria and denitrifying bacteria according to claim 1, characterized in that, The fermentation medium in S1 and S2 is the same, and the components are carbon source 5-30 g / L, nitrogen source 5-30 g / L, inorganic salt 1-10 g / L, metal ion 0-2 g / L, trace element 0.1-1 g / L, and the rest is water, pH 4-9.

0.

3. The method for treating high-salinity low-temperature wastewater with nitrifying bacteria and denitrifying bacteria according to claim 2, characterized in that, The carbon source is at least one of glucose, trisodium citrate, sodium pyruvate, sodium acetate and molasses.

4. The method for treating high-salinity low-temperature wastewater with nitrifying bacteria and denitrifying bacteria according to claim 2, characterized in that, The fermentation temperature of the nitrifying bacteria is 37 DEG C, and the fermentation time is 48 h; the fermentation temperature of the denitrifying bacteria is 37 DEG C, and the fermentation time is 50 h.

5. The method for treating high-salinity low-temperature wastewater with nitrifying bacteria and denitrifying bacteria according to claim 2, characterized in that, The nitrogen source is at least one of ammonium chloride, ammonium nitrate, sodium nitrate, ammonium sulfate, urea, beef extract, ammonia water, corn syrup and fish meal.

6. The method for treating high-salinity low-temperature wastewater with nitrifying bacteria and denitrifying bacteria according to claim 2, characterized in that, The inorganic salt is any one or combination of magnesium salt, nitrate, potassium salt, calcium salt, phosphate or hydrochloride.

7. The method for treating high-salinity low-temperature wastewater with nitrifying bacteria and denitrifying bacteria according to claim 2, characterized in that, The trace element is a mixture of ferrous sulfate 0.01-3 g / L, copper sulfate 0.001-1 g / L, zinc sulfate 0.01-0.5 g / L, calcium chloride 0.1-2 g / L, manganese sulfate 0.01-1 g / L and cobalt chloride 0.001-0.1 g / L.

8. The method for treating high-salinity low-temperature wastewater with nitrifying bacteria and denitrifying bacteria according to claim 1, characterized in that, The Alcaligenes FC-01052 is added first to treat the wastewater for 15 h, and then the Paracoccus denitrificans TD-20229 is added to treat the wastewater for 40 h.

9. The method for treating high-salinity low-temperature wastewater with nitrifying bacteria and denitrifying bacteria according to claim 8, characterized in that, The addition amount of the fermentation liquid to the wastewater to be treated is 2%-10%.

Images