CN107585880B - Method for treating high-salt phenol-containing wastewater by utilizing staphylococcus aureus enzyme preparation to strengthen microorganisms - Google Patents
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Abstract
The invention discloses a method for strengthening microbial treatment of high-salt phenol-containing wastewater by using a luteococcus enzyme preparation, which comprises the steps of domesticating activated sludge by using the enzyme preparation, and then treating the high-salt phenol-containing wastewater by combining the enzyme preparation with the domestication of the activated sludge, wherein the domestication speed of the activated sludge can be accelerated by using the enzyme preparation, the performance of the activated sludge can be improved, so that the efficiency of the microbial treatment of the wastewater can be strengthened, and the service life of a membrane can be prolonged.
Description
Technical Field
The invention belongs to the field of industrial wastewater treatment, and particularly relates to a method for treating high-salt phenol-containing wastewater by using a garcinia enzyme preparation reinforced microorganism.
Background
The phenolic substances belong to aromatic compounds, the main discharge types comprise phenol, chlorophenol and p-nitrophenol,is a poisonous and harmful pollutant which is difficult to degrade and is generated in chemical production. Due to the wide variety of phenolic compounds, which are difficult to degrade and have a "three-fold effect," 129 preferred pollutants and one of 65 toxic pollutants have been listed. In recent years, with the rapid development of industries such as petrochemical industry, pharmacy, building materials, pesticides, papermaking, coal gas, electronics, textiles, dyes, plastics, coking and the like, the discharge amount of phenol-containing wastewater is greatly increased, and the phenol-containing wastewater gradually becomes one of the main pollution sources of the current water body. Meanwhile, the phenol-containing wastewater has very complex components and often contains a large amount of salt (Na) besides organic pollution which is difficult to degrade+、Cl-、Ca+Etc.), when the phenol-containing wastewater with the total dissolved solid content of more than 3.5 percent is high-salt phenol-containing wastewater. The industrial wastewater containing high-salt and difficult-to-degrade organic pollutants not only causes serious pollution to the environment, but also increasingly harms human bodies, aquatic organisms and crops, and seriously damages ecological balance. For a long time, the treatment of high-salt phenol-containing wastewater is a difficult problem to be solved urgently in the field of sewage treatment.
Although phenol-containing wastewater treatment technologies based on physical and chemical principles, such as adsorption, extraction, membrane separation, advanced oxidation, etc., have been applied to a certain extent, the expected treatment effect is difficult to achieve due to the wide range of phenol types and contents in different wastewater, and the problems of low efficiency, high cost, secondary pollution, etc. in specific application. Because the microbial degradation treatment technology has the characteristics of safety, high efficiency, economy and the like, the technology becomes a research hotspot for the harmless treatment of the high-salt phenol-containing wastewater. However, the phenolic wastewater contains a large amount of compounds which are difficult to biodegrade and have biological toxicity, so that the growth and survival of microorganisms can be inhibited, for example, phenol has a remarkable inhibiting effect on related functional bacteria such as nitrogen and phosphorus removal in a biological treatment system, and meanwhile, the high-content salt in the phenolic wastewater inhibits the activity of the microorganisms in the biological treatment system, so that the traditional biological treatment method is difficult to achieve the treatment target. Researches show that the number of microorganisms which can be cultured by the traditional separation method only accounts for 1-15% of the total amount of the activated sludge microorganisms, and the industrial device for treating the phenol-containing wastewater contains abundant phenol hydroxylase gene types, while phenol-reducing bacteria with low encoding affinity constants cannot be obtained by the traditional enrichment culture method. A large number of potentially functional flora have not been investigated because they are in a viable but non-culturable (VBNC) state. In addition, most of the high-efficiency degradation strains or enrichment cultures obtained by laboratory separation enter a VBNC state under the stress of environmental pressure, so that the high-efficiency degradation strains or enrichment cultures show lower degradation activity in a biological system for actual wastewater treatment. How to improve the microbial activity of the biological treatment system of the high-salt phenol-containing wastewater to exert the optimal efficiency is a problem to be solved urgently in the biological treatment of the high-salt phenol-containing wastewater at present.
The discovery of the resuscitation-promoting factor (Rpf) of the staphylococcus luteus is considered as the most important breakthrough of resuscitation and culture of VBNC status bacteria. The garcinia Rpf is a secreted protein, can be secreted in an autocrine or paracrine mode, and has the molecular mass of 16-17 kDa. The mechanism of action of Rpf is generally accepted as Rpf, a lysozyme with peptidoglycan lytic enzyme activity, playing an important role in the cell wall lysis process. The method can not only recover and promote the growth of gram-positive (G +) bacteria with high G + C, but also has better recovery and promotion functions on G + bacteria with low G + C and part of gram-negative (G-) bacteria. Moreover, the staphylococcus luteus Rpf is used as a high-efficiency biological enhancer, the culturable number of bacteria can be increased by more than 100 times at the picomolar concentration, the growth of culturable cells can be promoted, and the promotion of potential functional flora in a wastewater biological treatment system by utilizing Rpf resuscitation has important significance. However, due to the limiting factors of complex extraction and purification of the Rpf protein, relatively high cost and the like, an economically feasible bioaugmentation method needs to be established. Research shows that the pure Rpf protein is easy to inactivate, and at the same time, at least two proteins with Rpf function (lysozyme activity) are separated from the supernatant of the garcinia lutescens, the molecular weights of the proteins are both larger than that of the Rpf protein, and the extracellular secretion of the garcinia lutescens contains at least 3 proteins with lysozyme activity. Therefore, the activity of the salt-tolerant phenol degrading flora can be recovered and promoted by using the garcinia enzyme preparation, the efficiency of the bioreactor is improved, and a wider separation bacterial source can be provided for culturing and screening high-efficiency salt-tolerant phenol degrading strains. Meanwhile, a new idea is provided for breaking the application bottleneck of the microbial treatment technology.
Disclosure of Invention
The invention aims to provide a method for treating high-salt phenol-containing wastewater by utilizing a staphylococcus aureus enzyme preparation to strengthen microorganisms aiming at the defects of low flora activity, slow growth speed and low degradation efficiency in the existing biological treatment system of the high-salt phenol-containing wastewater. The method has the characteristics of safety, low cost and high efficiency.
The purpose of the invention is realized by the following technical scheme: a method for strengthening microorganism treatment of high-salt phenol-containing wastewater by using a staphylococcus aureus enzyme preparation comprises the following steps:
(1) inoculating Flaccia luteus (Micrococcus luteus) into L MM culture medium, fermenting, culturing to obtain extracellular secretion, and further preparing 1-2U/m L activity of Flaccia luteus enzyme preparation;
(2) adding the enzyme preparation obtained in the step (1) into an inorganic salt culture medium with phenol as a unique carbon source according to the volume fraction of 3-4%, and acclimating the activated sludge;
(3) inoculating the acclimated sludge obtained in the step (2) into MBR according to the amount of 1-2 g/L M L SS, and adding enzyme preparation with volume fraction of 0.5-1.0% to treat high-salt phenol-containing wastewater.
Further, the step 1 specifically comprises:
(1.1) inoculating the staphylococcus luteus in L MM culture medium, culturing at 30 ℃ and 160r/min for 24-36h to obtain fermented seed liquid, inoculating the seed culture liquid into L MM culture liquid according to the inoculation amount of 2-4% (v/v), and culturing at 30 ℃ and 160r/min for 48-72h to obtain the staphylococcus luteus fermentation liquid;
the L MM medium comprises 4.0 g/L NH4Cl，1.4g/L KH2PO40.005 g/L biotin, 0.02 g/L L-methionine, 0.04 g/L vitamin B1, 1.0 g/L inosine, 0.03 g/L MgSO48.75 g/L L-lithium lactate salt, 1.5 ml/L mineral salt solution (0.375 g/L CuSO)4·5H2O，0.785g/L MnCl2·4H2O，0.18g/L FeSO4·7H2O，0.029g/L Na2MoO4·2H2O，0.089g/L ZnSO4·7H2O), pH 7.5;
(1.2) centrifuging the fermentation liquor obtained in the step 1.1 (8000-;
(1.3) putting the extracellular secretion into a 30KDa ultrafiltration centrifugal tube, centrifuging at 7500r/min for 10min, collecting liquid (liquid 1) at the lower part of an inner membrane of the tube, taking the liquid 1 to 10KDa ultrafiltration centrifugal tube, centrifuging at 7500r/min for 10min, collecting liquid (liquid 2) at the upper part of the inner membrane of the tube, adding solid ammonium sulfate into the liquid 2 to ensure that the mass fraction of the solid ammonium sulfate is 80% in a saturated state, stirring in an ice water bath for 1h, and centrifuging at 8000r/min for 20min to collect precipitates; and then adding phosphate buffer solution with the same volume as the liquid 2 into the precipitate, centrifuging the precipitate in a 10KDa ultrafiltration centrifugal tube at 7500r/min for 10min, and collecting the liquid on the upper part of the inner membrane of the tube, namely the luteolin enzyme preparation.
Compared with the existing method for treating high-salt phenol-containing wastewater by strengthening microorganisms, the method has the beneficial effects that:
1. the garcinia enzyme preparation used in the invention has the characteristics of low cost, simple and convenient operation, safety, no toxicity, green environmental protection and the like.
2. The invention can realize the high-efficiency domestication of the salt-tolerant phenol-reducing activated sludge by using the garcinia cambogia enzyme preparation, can recover the potential functional flora in the activated sludge, and can prevent the functional flora from entering a VBNC state, thereby shortening the domestication time of the activated sludge and playing the optimal salt-tolerant phenol-reducing performance of the activated sludge.
3. The optimum performance of indigenous microorganisms in the MBR can be maintained by utilizing the staphylococcus aureus enzyme preparation, the wastewater treatment time is shortened, the membrane pollution is reduced, and the service life of the membrane is prolonged, so that the wastewater treatment cost is reduced.
4. According to the invention, the luteolin enzyme preparation is added into the domestication culture medium of the salt-tolerant phenol-reducing activated sludge, and the comparison with a control group without the added enzyme preparation shows that the luteolin enzyme preparation can obviously improve the domestication efficiency of the salt-tolerant phenol-reducing activated sludge and promote the growth and the propagation of salt-tolerant phenol-reducing microorganisms.
5. According to the invention, the addition amount of the enzyme preparation in the activated sludge acclimation stage is 3-4% of the volume fraction, and the addition amount of the enzyme preparation in the MBR for wastewater treatment is only 0.5-1% of the volume fraction, so that the efficiency of activated sludge acclimation and wastewater treatment can be effectively improved, the characteristics of rapidness, sustainability, economy, practicability and the like are fully embodied, and the method has great popularization and application values.
6. The garcinia enzyme preparation disclosed by the invention not only can be used for rapidly domesticating salt-tolerant phenol-reducing flora and improving the efficiency of treating high-salt phenol-containing wastewater by microorganisms, but also can be used for promoting the recovery growth of functional flora in other industrial wastewater, and provides a rapid, efficient, safe, environment-friendly and low-cost method for applying the microbial technology to the biological treatment of refractory organic wastewater and composite polluted wastewater.
In conclusion, the method accords with the environmental protection concept of green safety, realizes the reinforced microbial treatment of the difficultly degraded and compound polluted wastewater, fully and effectively exerts the degradation performance of indigenous microbes in the wastewater biological treatment system, and provides a safe, cheap and efficient method for promoting the popularization and application of the microbial technology for treating the complex industrial wastewater.
Drawings
FIG. 1 is a graph of the effect of adding a enzyme preparation of Garcinia cambogia on the salt tolerance of activated sludge;
FIG. 2 is a graph showing the effect of adding a Garcinia cambogia enzyme preparation on the phenol resistance of activated sludge.
Detailed Description
The invention relates to a method for strengthening microbial treatment of high-salt phenol-containing wastewater by using a staphylococcus aureus enzyme preparation, which comprises the following steps of:
1. preparation of enzyme preparation of Garcinia cambogia
Activation culture of Garcinia Cambogia, the Garcinia Cambogia (Micrococcus L uteus IAM14879) was purchased from the Japan institute of physical and chemical engineering, microorganism culture Collection, L MM medium (4.0 g/L NH)4Cl，1.4g/L KH2PO40.005 g/L biotin, 0.02 g/L L-methionine, 0.04 g/L vitamin B1, 1.0 g/L inosine, 0.03 g/L MgSO48.75 g/L L-lithium lactate salt, 1.5 ml/L mineral salt solution (0.375 g/L CuSO)4·5H2O，0.785g/L MnCl2·4H2O，0.18g/L FeSO4·7H2O，0.029g/L Na2MoO4·2H2O，0.089g/L ZnSO4·7H2O), pH 7.5) into ampoules containing the powder of luteus, left at room temperature for 20min and streaked with an inoculating loop to L MM solid plates.
And (3) performing fermentation culture on the garcinia lutescens, namely selecting 1 ring of the garcinia lutescens on an L MM solid plate, inoculating the garcinia lutescens into a 100m L triangular flask with a 20m L L MM liquid culture medium, culturing for 32h at 30 ℃ at 160r/min, then inoculating the seed culture solution into a L MM culture solution according to the inoculation amount of 3% (v/v), and culturing for 52h at 30 ℃ at 160r/min to obtain the garcinia lutescens fermentation liquor.
Preparation of a Garcinia cambogia enzyme preparation: centrifuging the fermentation liquid of Garcinia lutescens at 8000r/min for 15min to remove thallus, and filtering with 0.22 μm filter membrane to obtain extracellular secretion of Garcinia lutescens. Taking a certain volume of extracellular secretion to a 30KDa ultrafiltration centrifugal tube, centrifuging at 7500r/min for 10min, collecting liquid (liquid 1) at the lower part of an inner membrane of the tube, then taking the liquid 1 to 10KDa ultrafiltration centrifugal tube, centrifuging at 7500r/min for 10min, collecting liquid (liquid 2) at the upper part of the inner membrane of the tube, adding a certain amount of solid ammonium sulfate into the liquid 2 to ensure that the mass fraction of the solid ammonium sulfate is 80% in a saturated state, stirring in an ice water bath for 1h, centrifuging at 8000r/min for 20min, and collecting precipitates; and then adding phosphate buffer solution with the same volume as the liquid 2 into the precipitate, centrifuging the precipitate in a 10KDa ultrafiltration centrifugal tube at 7500r/min for 10min to collect the liquid on the upper part of the inner membrane of the tube, namely the taffia glomerata enzyme preparation, and storing the liquid at the temperature of minus 20 ℃ for later use.
2. Detection of activity of enzyme preparation of Garcinia cambogia
Drawing a glucosamine standard curve, namely respectively taking 0, 0.5, 1.0, 1.5 and 2.0m L of a glucosamine standard solution (with the concentration of 50 mu g/ml) into a colorimetric tube, respectively adding distilled water to 5m L, then adding a 1m L solution of an acetylacetone solution, placing the colorimetric tube in a boiling water bath for reaction for 25min, rapidly cooling the colorimetric tube by using ice, then adding 3m L absolute ethyl alcohol and 1m L p-dimethylaminobenzaldehyde, strongly oscillating the colorimetric tube, standing the colorimetric tube at 20-25 ℃ for 1h, taking the non-glucosamine standard solution tube as a blank control, and using A of the glucosamine standard solution with other content527And drawing a standard curve. The standard curve of glucosamine obtained isY＝0.247X-0.0418(R2＝0.970)。
The activity of the garcinia preparation is measured by adding 2mg of peptidoglycan into 15 mu L of garcinia preparation (test group) and distilled water (blank group), respectively, after 2 hours of action at 30 ℃, transferring the two groups into 10m L volumetric flasks, adding 1.5m L HCl with concentration of 6mol/m L, hydrolyzing for 1 hour in boiling water bath, after cooling, neutralizing to pH 7.0 with sodium hydroxide, diluting to scale with water, taking 2m L, adding 2m L distilled water, adding 1m L acetylacetone, after 25 minutes of reaction in boiling water bath, rapidly cooling with ice, adding 3m L anhydrous ethanol and 1m L p-dimethylaminobenzaldehyde, strongly shaking, standing for 1 hour at 20-25 ℃, measuring absorbance values of 0.089 and 0.112 at A527 for the test group and the blank group respectively, calculating glucosamine)/(content of 26.5 mu)/(g and 31.13 g respectively by standard curve, reducing glucosamine activity by weight of 26.13 g, and obtaining a glucosamine reaction sample with weight of 30g (30 m 4642) and weight of 30.38 g, and reducing glucosamine by weight of 30g (30 m 4642) through dilution of a reaction time of 30.36 g (30 m L) and 30 m 464. multidot-3) 1.116U/m L, the activity of the obtained enzyme preparation of Garcinia cambogia was 1.116U/m L.
3. Domestication and culture of efficient salt-tolerant phenol-reducing activated sludge
Acclimatization of activated sludge, namely adding 3% of a luteolongin enzyme preparation by volume fraction into an inorganic salt culture medium taking phenol as a unique carbon source to obtain an acclimatized culture medium of a treatment group, wherein the composition of the inorganic salt culture medium is 0.5 g/L KH2PO4，0.5g/L K2HPO4，0.2g/L MgSO4，1g/L NH4Cl, trace salt solution 10m L/L (4 mg/L MoO)3，28mg/LZnSO4·5H2O，0.02mg/L CuSO4·5H2O，4mg/L H3BO3，4mg/L MnSO4·5H2O，4mg/L CoCl2·6H2O), phenol and NaCl concentrations were determined according to the experimental requirements, with a pH of 7.0. The control group was acclimatized in inorganic salt medium with phenol as the sole carbon source without addition of enzyme preparation.
The acclimation process of the activated sludge is divided into four stages, wherein the phenol concentration is continuously increased according to the gradient of 200 mg/L, 500 mg/L, 1000 mg/L and 1500 mg/L, the salinity load is synchronously increased at the same time, the NaCl gradient of 5 g/L, 10 g/L, 15 g/L and 30 g/L is gradually increased, the phenol concentration in the supernatant of the muddy water is periodically measured, the change of activated sludge flocs and the color is observed, in different acclimation stages, when the phenol removal rate reaches 100%, the supernatant is discharged after the activated sludge is balanced for 2d, a fresh culture medium is replaced, the phenol concentration and the salinity are improved, and then the acclimation is carried out in the next stage, and the time required by the treatment group and the control group for respectively acclimating to obtain the activated sludge No. 1 and No. 2 which can completely degrade 1500 mg/L phenol under the salinity of 30 g/L is respectively 65.
4. Influence of the luteolin enzyme preparation on salt tolerance and phenol tolerance of activated sludge
The influence of the enzyme preparation on the salt tolerance of the activated sludge is that the two groups of domesticated activated sludge are respectively inoculated into synthetic wastewater with the same phenol concentration (1500 mg/L) and different salt contents (20, 60, 100, 140 and 180 g/L) according to the inoculation amount of 2 g/L, the phenol residue in the wastewater is respectively measured after the synthetic wastewater is cultured for 100 hours at the temperature of 30 ℃ and 180r/min at 160-.
The influence of the enzyme preparation on the phenol resistance of the activated sludge is that the two groups of domesticated activated sludge are respectively inoculated into synthetic wastewater containing the same salt concentration (30 g/L) and different phenol concentrations (1500, 2000, 2500, 3000, 3500 and 4000 mg/L) according to the inoculation amount of 2 g/L, the temperature is 30 ℃, the temperature is 160-.
Measuring phenol content by 4-amino-substituted biao method, which comprises adding 100 μ L sample into 100 μ L buffer solution, 200 μ L4-amino-substituted biao solution, and 200 μ L potassium ferricyanide solution, diluting to 10m L, standing for 15min, measuring absorbance at 510nm with ultraviolet spectrophotometer, and drawing standard curvePrecisely measuring phenol standard solution 5.0, 7.5, 10.0, 12.5, 15.0, 17.5 and 20.0m L-50 m L volumetric flasks, adding 1.0m L buffer solution (20 g of ammonium chloride NH4Cl is dissolved in 100m L ammonia water), mixing uniformly, sequentially adding 4-aminoantipyrine solution 2.0m L, potassium ferricyanide solution 2.0m L, redistilled water to a constant volume of 50m L, and measuring the absorbance of different phenol concentrations at a wavelength of 510nm, wherein the obtained phenol standard curve is Y0.1373X +0.0009(R is 0.1373X + 0.0009)20.9998). The measured absorbance is substituted into the standard curve to obtain the phenol concentration, thereby calculating the removal rate of phenol.
5. Influence of a luteococcus enzyme preparation on the structure of acclimatized activated sludge flora
The two groups of domesticated activated sludge samples No. 1 (added with enzyme preparation) and No. 2 (not added with enzyme preparation) are subjected to high-throughput sequencing analysis of 16SrRNA gene. FastDNA is adopted as DNA of an activated sludge sampleTMThe method comprises the steps of extracting a 16S rRNA gene V3-V4 hypervariable region fragment by using a Kit SPIN Kit for Soil (Bio101Inc., USA), amplifying 16S rRNA gene V3-V4 hypervariable region fragments by using primers 341F5 '-CCTAYGGGRBGCASCAG-3' and 806R 5 '-GGACTACNNGGGTATCTAAT-3', performing RCR reaction system of 25 mu L as shown in 3.2.3.1, performing PCR reaction under the conditions of pre-denaturation at 94 ℃ for 2min, denaturation at 94 ℃ for 30S, annealing at 57 ℃ for 30S, extension at 72 ℃ for 30S, performing 25 cycles, final extension at 72 ℃ for 5min, and keeping at 16 ℃, mixing three groups of parallel PCR products of each sample in equal amount, performing double-end sequencing by using Illumina Miseq 2500 platform after quantitative and equal quality control, then performing operational classification unit (OTU) on effective sequences by using Uprase software, clustering the sequences into OTOI bacteria according to 97% similarity, performing OTNOSE group analysis on different classification levels, and using a preparation for promoting the growth of microorganisms of Bacteromycota strain group, wherein the growth of the strain can be changed as shown in the phylum of Bacillus.
TABLE 1 relative abundance of different phyla
| Phylum (relative abundance%) | Raw sludge | Treatment group-activated sludge | Control group-activated sludge |
| Proteobacteria | 77.95 | 47.39 | 59.56 |
| Bacteroidetes | 12.27 | 42.01 | 31.06 |
| Actinobacteria | 2.64 | 6.17 | 3.41 |
| Firmicutes | 4.39 | 1.60 | 2.11 |
| Spirochaetes | 0.65 | 0.73 | 0.92 |
| Tenericutes | 0.39 | 0.30 | 0.36 |
| Chrysiogenetes | 0.23 | 0.23 | 0.14 |
| Synergistetes | 0.20 | 0.19 | 0.10 |
| Saccharibacteria | 0.16 | 0.16 | 0.15 |
| Others | 1.12 | 1.22 | 2.19 |
6. Efficiency of enzyme preparation enhanced MBR (membrane bioreactor) for treating high-salt phenol-containing wastewater
Respectively inoculating the domesticated activated sludge sample No. 1 (added with an enzyme preparation) and No. 2 (not added with the enzyme preparation) to 4 groups of same Membrane Bioreactors (MBRs) according to the amount that M L SS is 1 g/L, adding a tawnia glomerata enzyme preparation with volume fraction (v/v) of 0.8% into a treatment group, and adding no enzyme preparation into a control group to obtain 4 sets of devices for treating high-salt phenol-containing wastewater:
set 1: activated sludge No. 1 + MBR + adding 0.8% of enzyme preparation;
and (2) set: activated sludge No. 1 + MBR + without 0.8% enzyme preparation;
set 3: activated sludge No. 2 + MBR + adding 0.8% of enzyme preparation;
set 4: activated sludge No. 2 + MBR + no 0.8% enzyme preparation added;
treating wastewater containing 30 g/L NaCl, 1000 mg/L phenol, 2450 mg/L COD and 212.5 mg/L ammonia nitrogen, controlling the temperature at 30 ℃, the pH at 8, the HRT at 24h and the inflow rate at 6m L/min;
the concentration of the phenol in the effluent of the 1 st set is 0 mg/L of 46 mg/L, the concentration of ammonia nitrogen is 0.8 mg/L0, the concentration of the phenol in the effluent of the 2 nd set is 56 mg/L1, COD is 145 mg/L, the concentration of the ammonia nitrogen is 7.2 mg/L, the concentration of the phenol in the effluent of the 3 rd set is 93 mg/L of 237 mg/L, the concentration of the ammonia nitrogen is 10.6 mg/L, the concentration of the phenol in the effluent of the 4 th set is 124 mg/L of 302 mg/L, the concentration of the ammonia nitrogen is 13.7 mg/L, compared with the 1 st set and the 2 th set, the TMP is reduced by 9KPa, compared with the 1 st set and the 4 th set, the TMP is reduced by 13KPa, the period of the membrane needing to be cleaned is 25d, the 2 nd set is 14d, the 3 rd set is 11d, and the MBR is 7d, the result shows that the membrane pollution speed can be obviously reduced by adding the enzyme preparation, the service life of the membrane pollution can be obviously prolonged, and the high-effect of the tequillaena wastewater treatment device can be obviously enhanced by.
In addition, the inoculation amount of the domesticated sludge is ensured to be 1-2 g/L, the addition amount of the enzyme preparation is 0.5-1.0 vol%, and the high-efficiency wastewater primary draft treatment capability can be realized.
Claims (2)
1. A method for strengthening microorganism treatment of high-salt phenol-containing wastewater by using a staphylococcus aureus enzyme preparation is characterized by comprising the following steps:
(1) inoculating Flaccia luteus (Micrococcus luteus) into L MM culture medium, fermenting, culturing to obtain extracellular secretion, and further preparing 1-2U/m L activity of Flaccia luteus enzyme preparation;
(2) adding the enzyme preparation obtained in the step (1) into an inorganic salt culture medium with phenol as a unique carbon source according to the volume fraction of 3-4%, and acclimating the activated sludge;
(3) inoculating the acclimated sludge obtained in the step (2) into MBR according to the amount of 1-2 g/L M L SS, and adding enzyme preparation with volume fraction of 0.5-1.0% to treat high-salt phenol-containing wastewater.
2. The method according to claim 1, characterized in that step (1) is in particular:
(1.1) inoculating the garcinia to L MM culture medium, culturing at 30 ℃ and 160r/min for 24-36h to obtain fermented seed liquid, inoculating the fermented seed liquid into L MM culture liquid according to the inoculation amount of 2-4% (v/v), and culturing at 30 ℃ and 160r/min for 48-72h to obtain the garcinia fermentation liquid;
the L MM medium comprises 4.0 g/L NH4Cl，1.4g/L KH2PO40.005 g/L biotin, 0.02 g/L L-methionine, 0.04 g/L vitamin B1, 1.0 g/L inosine, 0.03 g/L MgSO48.75 g/L L-lithium lactate, 1.5 ml/L mineral salt solution including 0.375 g/L CuSO4·5H2O，0.785g/L MnCl2·4H2O，0.18g/L FeSO4·7H2O，0.029g/L Na2MoO4·2H2O，0.089g/L ZnSO4·7H2O, pH 7.5;
(1.2) centrifuging the fermentation liquor obtained in the step (1.1) for 10-15min at 8000-10000r/min to remove thalli, and performing sterile filtration by a 0.22 mu m filter membrane to obtain an extracellular secretion of the staphylococcus aureus;
(1.3) transferring the extracellular secretion to a 30KDa ultrafiltration centrifugal tube, centrifuging at 7500r/min for 10min, collecting the liquid on the lower part of the inner membrane of the tube, marking as liquid 1, then taking the liquid 1-10 KDa ultrafiltration centrifugal tube, centrifuging at 7500r/min for 10min, collecting the liquid on the upper part of the inner membrane of the tube, marking as liquid 2, adding solid ammonium sulfate into liquid 2 to ensure that the mass fraction of the solid ammonium sulfate is 80% in a saturated state, stirring in an ice water bath for 1h, centrifuging at 8000r/min for 20min, and collecting precipitates; and then adding phosphate buffer solution with the same volume as the liquid 2 into the precipitate, centrifuging the precipitate in a 10KDa ultrafiltration centrifugal tube at 7500r/min for 10min, and collecting the liquid on the upper part of the inner membrane of the tube, namely the luteolin enzyme preparation.
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