CN103060420A - Interaction of polyphenol pollutants in the process of biodegradation and a research method thereof - Google Patents
Interaction of polyphenol pollutants in the process of biodegradation and a research method thereof Download PDFInfo
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Abstract
The present invention relates to interaction of polyphenol pollutants in the process of biodegradation and a research method thereof. The research method comprises: preparing medium for degrading polyphenol pollutants; adding phenol or cresol into mineral medium as a carbon source to form the selective medium for phenol degradation experiments; in the study of influence of different m-cresol concentrations to phenol degradation, fixing the concentration of phenol, and selecting several gradient levels of different m-cresol concentrations; in the study of influence of different phenol concentrations to m-cresol degradation by using Pseudomonas aeruginosa GIMT1.074 inoculating into the medium, fixing the concentration of m-cresol, and selecting several gradient levels of different phenol concentrations; performing parallel experiments to each gradient; and analyzing changes of cell concentrations in the culture process, and the changes of degradation of phenol and cresol, to obtain interaction of polyphenol pollutants in the process of biodegradation; which has some theoretical guiding significance and practical value for biodegradation of phenol pollutants in the actual environment.
Description
Technical field
The present invention relates to interaction and research method in a kind of polyphenol pollutent biodegradation process.
Background technology
Phenolic compound refers to the general designation of a series of aromatic hydrocarbons hydroxy derivatives, can seriously stimulate respiratory system and eyes, corrosion skin causes burns and dermatitis, even cause chronic poisoning to cause the Digestive tract obstacle, neurological disorder, internal organ are impaired etc., can it be solidified with protein bound at cell interior, thereby cell killing has the toxicity hazard of very big protoplasma.The phenolic compound oxygen-consumption that exists in water is higher, has seriously broken the oxygen balance in the water ecology, and (1-10mg/L) can cause the mortality of water body biology under low concentration, suppresses various microorganism growth, destroys the water ecology balance.Containing the waste water of phenolic comp ' ds pollution mainly from chemical enterprises such as petroleum chemical plant, plastic molding and processing plant, Fibre Plant, refinery, resin processing plant and coke-oven plants, is one of important pollutent of water body.Phenolic comp ' ds pollution all can produce toxicity to all biologically active bodies, can be by directly entering blood circulation with contacting without liver detoxification of skin, mucous membrane, and cause cytoclasis and lose vigor, also can invade human body by the oral cavity, cause cell injury.The phenol liquid of high density can make protein coagulating, and can continue to the body inner penetration, causes the deep tissue damage, and necrosis and even whole body are poisoned, even the phenol liquid of lower concentration also can make protein denaturation.If people's long-term drinking can be caused chronic poisoning by the water that phenol pollutes, anaemia, giddy occur, be losing one's memory and various neural disease, serious meeting causes death.Contain simultaneously the multiple typical difficult degradation phenolic compounds such as phenol, cresols, chlorophenol, nitrophenol in the industry phenolic wastewater, realize that its harmless treatment is a global problem all the time.Phenolic compound is classified as one of 129 kinds of priority pollutants by American National environment protection general bureau (EPA), and the phenol concentration in the regulation waste water must not surpass lmg/L.China also lists it in " priority pollutants Black List in the water " simultaneously, and the discharging of phenolic wastewater is had strict standard.
The processing means of phenolic comp ' ds pollution have Physical, chemical method and biological process.Biological process degraded organic industrial sewage wherein is to use at present the most general Treatment of Phenol Containing Water, but in biological treatment process, organic pollutant causes its degradation efficiency low to the toxic action of microorganism, and obtaining specific microbial strains becomes the key problem that realizes the pollutent efficient degradation.But processing from the laboratory lab scale to actual waste water the application process, often because the complicacy of pollutent and refractory organics cause the process collapse.Research finds that there are some peculiar microorganism floras in occurring in nature, because the diversity of its system's constitute and function can show the organic pollutant degradation metabolic capacity that single strain does not possess.Under laboratory condition, the mixed culture of multiple pure culture microorganism also shows the raising to the Degradation of a certain or several phenolic comp ' ds pollution, even the degradation capability that can obtain originally not have.High efficiency and stability under the high pollution thing pressure that flora constitute and function diversity is brought are for phenolic comp ' ds pollution high-performance bio degraded provides new approaches after active sludge and pure culture bacterial classification.At present, the improvement method of phenolic wastewater progressively grows up along with phenols is endangered understanding and water technology development, mainly is divided into three major types: physical treatment process, method of chemical treatment and biological treatment.And along with technology constantly develops the continuous combination of the whole bag of tricks and interpenetrates, to reach better regulation effect.Biological process degraded organic industrial sewage is to use at present the most general Treatment of Phenol Containing Water, the so far history in existing more than 100 year.Its principle of work is to utilize the metabolism of microorganism that Organic Pollutants in Wastewater is converted into CO
2, N
2And H
2The discharging of the toxicological harmless small-molecule substances such as O.The advantages such as the biological process processing efficiency is high, it is wide to use, processing power is strong, equipment is simple, produce that secondary pollution is few, effluent quality good, operation and easy to operate and working cost are lower are occupied an leading position in the current industrial wastewater processing technology.But it is larger that biological treatment process is subjected to waste water ph, temperature and contains the impact of the factors such as phenol amount and kind, so require stricter to operational condition.Although a large amount of phenol degrading wild mushrooms is out screened, phenolic wastewater often contains multiple difficult degradation compound simultaneously in producing, and limits the application development of pure bacterial strain.Recent researches finds that the co-cultivation of multiple-microorganism is conducive to the combination of their metabolic functions.Often have multiple pollutant in actual waste water, exist various interactions in its degradation process, main manifestations is the common metabolic patterns of mutual inhibition and promotion.Therefore single substrate model can not be described the degraded of this class mixture.
Because complicacy and the diversity of phenolic comp ' ds pollution, so various components have important practical significance to the effect of microbiological deterioration in the research phenolic comp ' ds pollution.
Summary of the invention
The invention provides the interaction in a kind of polyphenol pollutent biodegradation process.Another object of the present invention provides this interactional a kind of research method.The phenolic comp ' ds pollution degradation bacteria that the present invention is applied to is Pseudomonas aeruginosa GIMT1.074.
Concrete technical scheme is as follows:
Interactional research method in a kind of polyphenol pollutent biodegradation process, its step is as follows:
1) configuration is used for the substratum of degraded polyphenol pollutent; The used selective medium of phenols degradation experiment is to add phenol or meta-cresol on the minimal medium basis as carbon source; At the different meta-cresol concentration Degradation of Phenols of research when affect, the fixing concentration of phenol, several gradient levels of the concentration different concns of selection meta-cresol; When the different phenol concentration of research affects the degraded of meta-cresol, the fixing concentration of meta-cresol, several gradient levels of the different concns of selection phenol; Each gradient is carried out parallel laboratory test;
2) utilizing Pseudomonas aeruginosa GIMT1.074 to be inoculated in the substratum cultivates;
3) adopt optical densitometric method and constant weight desiccating method to measure cell concentration; Adopt the aldehydes matter concentration in the high effective liquid chromatography for measuring degraded system;
4) according to the variation of cell concn in the culturing process and the variation of phenol and meta-cresol degraded situation, obtain the interaction data in the polyphenol pollutent biodegradation process.
Described substratum is: 500mg (NH
4)
2SO
4, 200mg KH
2PO
4, 100mg MgSO
4, 800mgNa
2HPO
412H
2O, the 20mg yeast soaks powder, 10mL trace elements mother liquor and 1000mL H
2O.
Described trace elements mother liquor composition is as follows: 0.4g MnSO
44H
2O, 0.4g ZnSO
47H
2O, 0.1gNa
2MoO
42H
2O, 0.1g CuSO
45H
2O, 1.0g CaCl
2, 10.0g Na
2SO
4, 2.0g FeSO
47H
2O, 0.5mL H
2SO
4, 2.0g NaOH, 12.0g disodium ethylene diamine tetraacetate and 1000mL H
2O.
Described step 2) cultural method is: with pure initial OD
600Value is that 0.01 Pseudomonas aeruginosa GIMT1.074 is inoculated in the substratum, and all initial pH value of medium are 7.2, and liquid culture and solid culture temperature are 30 ℃, and rotating speed was 200rpm when shaking table was cultivated, and incubation time is 40 hours.
Described optical densitometric method is measured the cell concentration method: optical density(OD) is done blank with corresponding substratum, measures the absorbancy (OD of bacteria suspension under the 600nm wavelength
600); The concentration of sample need to be diluted to multiply each other with measured result and extension rate below 0.8 when surpassing 0.8 again and obtain the actual OD of bacterium liquid
600Value.
6. the method for claim 1 is characterized in that constant weight dry method measurement cell concentration method is: to dry weight, the bacterium liquid of certain volume physiological period different concns got in the weighing record with resistant to elevated temperatures centrifuge tube constant weight, and centrifugal, supernatant liquor inclines; Wash cell with sterile distilled water, repetitive scrubbing twice is abandoned supernatant, then will be dried to constant weight, the weighing record in centrifuge tube and the thalline placement loft drier.
Described high performance liquid chromatography is: high performance liquid chromatography chromatogram model is Agilent high performance liquid chromatography 1200, and the chromatographic column model is Eclipse XDB-C18,4.6 * 250mm, 5 μ m; Moving phase adopts volume ratio methyl alcohol: water=4:3, flow velocity 1.0mL/min; Sample size is 10 μ L, 30 ℃ of column temperatures, and the UV device detects wavelength 280nm.
The present invention utilizes in the process of Pseudomonas aeruginosa GIMT1.074 degradation of phenol and meta-cresol mixture and finds:
1) restraining effect of the higher Degradation of Phenol of concentration of meta-cresol is larger, and it is longer that cell concn reaches the used time of same OD value, but after the reaction times of abundance, meta-cresol concentration is higher, and final cell concentration is larger; Simultaneously, the concentration of meta-cresol is higher, and the needed time of degradable phenol and meta-cresol is longer;
2) concentration of phenol is higher, and degraded has certain restraining effect to meta-cresol, and it is shorter that cell concn reaches the used time of same OD value, but after the reaction times of abundance, phenol concentration is higher, and final cell concentration is larger; Simultaneously, the concentration of phenol is higher, and the degradation time of degradable meta-cresol and phenol is longer.
The present invention analyzes the variation of cell concn in the culturing process and the variation of phenol and meta-cresol degraded situation, obtain the interaction in the polyphenol pollutent biodegradation process, this biological degradation for phenolic comp ' ds pollution in the actual environment has certain theory directive significance and actual application value.
Description of drawings
Fig. 1 is that the meta-cresol of different concns exists the impact on Pseudomonas aeruginosa GIMT1.074 phenol biodegradation characteristic, comprising the biomass variation tendency of thalline, phenol concentration in the system change and the degraded system in the change in concentration of meta-cresol.
Fig. 2 is that the phenol of different concns exists the impact on Pseudomonas aeruginosa GIMT1.074 meta-cresol biodegradation character, comprising the biomass variation tendency of thalline, phenol concentration in the system change and the degraded system in the change in concentration of meta-cresol.
Embodiment
By will helping further to understand the present invention below in conjunction with specific examples, but protection scope of the present invention is not restricted to this:
1 configuration is used for the substratum of degraded polyphenol pollutent:
The composition of A, minimal medium is (NH
4)
2SO
4500mg, KH
2PO
4200mg, MgSO
4100mg, Na
2HPO
412H
2O 800mg, yeast soak powder 20mg, trace elements mother liquor 10mL, H
2O 1000mL; Wherein the trace elements mother liquor composition is as follows: MnSO
44H
2O 0.4g, ZnSO
47H
2O 0.4g, Na
2MoO
42H
2O, 0.1g, CuSO
45H
2O 0.1g, CaCl
21.0g, Na2SO
410.0g, FeSO
47H
2O 2.0g, H
2SO
40.5mL, NaOH 2.0g, disodium ethylene diamine tetraacetate 12.0g, H
2O 1000mL.
B, the used selective medium of phenols degradation experiment are to add phenol or meta-cresol on the minimal medium basis as carbon source.The phenol sterilization is through 0.22 μ m membrane filtration, and other reagent and medium sterilization carry out 30min under 115 ℃.When affect, fixedly the concentration of phenol is 600mg/L at the different meta-cresol concentration Degradation of Phenols of research, and the concentration of selection meta-cresol is 100,200,300,400, five gradient levels of 500mg/L.When the different phenol concentration of research affect the degraded of meta-cresol, fixedly the concentration of meta-cresol is 400mg/L, the concentration of selecting phenol is 200,300,400,500, five gradient levels of 600mg/L, cultivates 40 hours in the shaking table by the rotating speed under 200rpm and 30 ℃; Ten gradients altogether, each gradient is carried out three groups of parallel laboratory tests.。Cell concentration in per 5 hours sampling and measuring reaction systems and the concentration of remaining phenolic comp ' ds pollution are also analyzed, and association has obtained the influence mode of heterogeneity to the phenolic comp ' ds pollution degraded.
2 Pseudomonas aeruginosa GIMT1.074 are inoculated in the substratum and cultivate:
With pure initial OD
600Value is that 0.01 Pseudomonas aeruginosa GIMT1.074 is inoculated in the substratum, cultivates, and culture condition is as follows: all initial pH value of medium are 7.2, and liquid culture and solid culture temperature are 30 ℃, and rotating speed was 200rpm when shaking table was cultivated.Incubation time is 40 hours.
Cell concn in 3 degradation processes and aldehydes matter method for measurement of concentration are:
A. the sample that took out in the substratum every 5 hours is measured the concentration of thalline and the aldehydes matter concentration in the degraded system, and the method for measuring cell concentration is as follows:
The a cell concentration adopts optical densitometric method and constant weight desiccating method to measure.Concrete operations are as follows: optical density(OD) is done blank with corresponding substratum, measures the absorbancy (OD of bacteria suspension under the 600nm wavelength
600).The concentration that it should be noted that sample need to be diluted to multiply each other with measured result and extension rate below 0.8 when surpassing 0.8 again and obtain the actual OD of bacterium liquid
600Value.
B constant weight desiccating method, to dry weight, the bacterium liquid of certain volume physiological period different concns got in the weighing record with resistant to elevated temperatures centrifuge tube constant weight, and centrifugal, supernatant liquor inclines.Wash cell with sterile distilled water, repetitive scrubbing twice is abandoned supernatant, then will be dried to constant weight, the weighing record in centrifuge tube and the thalline placement loft drier.
B. the aldehydes matter method for measurement of concentration in the degraded system is:
The concentration of aldehydes matter adopts high performance liquid chromatography (HPLC) method to measure in the nutrient solution.The chromatogram model is Agilent (Agilent) high performance liquid chromatography 1200, and the chromatographic column model is Eclipse XDB-C18,4.6 * 250mm, 5 μ m.Moving phase adopts methyl alcohol: water=4:3(V:V, flow velocity 1.0mL/min.Sample size is 10 μ L, 30 ℃ of column temperatures, and the UV device detects wavelength 280nm.
Concrete test data is as follows:
Embodiment 1
Phenol concentration is on the impact of meta-cresol degraded
Dispose on request the minimal medium of phenols degraded, under 115 ℃, carry out the 30min sterilization.The fixing meta-cresol that adds 400mg/L, and the phenol of 200mg/L, cultivated 40 hours in the shaking table of the rotating speed under 200rpm and 30 ℃, the concentration that recorded cell with the dry cell weight method of masurement at the 10th hour is 86.31mg/L, uses phenol that liquid chromatograph measures and the concentration of meta-cresol to be respectively 100.45mg/L and 348.79mg/L.
Embodiment 2
Phenol concentration is on the impact of meta-cresol degraded
Dispose on request the minimal medium of phenols degraded, under 115 ℃, carry out the 30min sterilization.The fixing meta-cresol that adds 400mg/L, and the phenol of 200mg/L, cultivated 40 hours in the shaking table of the rotating speed under 200rpm and 30 ℃, the concentration that recorded cell with the dry cell weight method of masurement at the 15th hour is 151.34mg/L, uses phenol that liquid chromatograph measures and the concentration of meta-cresol to be respectively 3.01mg/L and 276.34mg/L.
Embodiment 3
Phenol concentration is on the impact of meta-cresol degraded
Dispose on request the minimal medium of phenols degraded, under 115 ℃, carry out the 30min sterilization.The fixing meta-cresol that adds 400mg/L, and the phenol of 200mg/L, cultivated 40 hours in the shaking table of the rotating speed under 200rpm and 30 ℃, the concentration that recorded cell with the dry cell weight method of masurement at the 20th hour is 213.51mg/L, uses phenol that liquid chromatograph measures and the concentration of meta-cresol to be respectively 0mg/L and 75.24mg/L.
Embodiment 4
Phenol concentration is on the impact of meta-cresol degraded
Dispose on request the minimal medium of phenols degraded, under 115 ℃, carry out the 30min sterilization.The fixing meta-cresol that adds 400mg/L, and the phenol of 200mg/L, cultivated 40 hours in the shaking table of the rotating speed under 200rpm and 30 ℃, the concentration that recorded cell with the dry cell weight method of masurement at the 25th hour is 299.61mg/L, uses phenol that liquid chromatograph measures and the concentration of meta-cresol to be respectively 0mg/L and 5.49mg/L.
Embodiment 5
Phenol concentration is on the impact of meta-cresol degraded
Dispose on request the minimal medium of phenols degraded, under 115 ℃, carry out the 30min sterilization.The fixing meta-cresol that adds 400mg/L, and the phenol of 600mg/L, cultivated 40 hours in the shaking table of the rotating speed under 200rpm and 30 ℃, the concentration that recorded cell with the dry cell weight method of masurement at the 10th hour is 69.28mg/L, uses phenol that liquid chromatograph measures and the concentration of meta-cresol to be respectively 596.96mg/L and 362.49mg/L.
Embodiment 6
Phenol concentration is on the impact of meta-cresol degraded
Dispose on request the minimal medium of phenols degraded, under 115 ℃, carry out the 30min sterilization.The fixing meta-cresol that adds 400mg/L, and the phenol of 600mg/L, cultivated 40 hours in the shaking table of the rotating speed under 200rpm and 30 ℃, the concentration that recorded cell with the dry cell weight method of masurement at the 20th hour is 326.95mg/L, uses phenol that liquid chromatograph measures and the concentration of meta-cresol to be respectively 286.45mg/L and 321.41mg/L.
Embodiment 7
Phenol concentration is on the impact of meta-cresol degraded
Dispose on request the minimal medium of phenols degraded, under 115 ℃, carry out the 30min sterilization.The fixing meta-cresol that adds 400mg/L, and the phenol of 600mg/L, cultivated 40 hours in the shaking table of the rotating speed under 200rpm and 30 ℃, the concentration that recorded cell with the dry cell weight method of masurement at the 30th hour is 658.34mg/L, uses phenol that liquid chromatograph measures and the concentration of meta-cresol to be respectively 0mg/L and 103.81mg/L.
Embodiment 8
The impact of meta-cresol concentration Degradation of Phenol
Dispose on request the minimal medium of phenols degraded, under 115 ℃, carry out the 30min sterilization.The fixing phenol that adds 600mg/L, and the meta-cresol of 100mg/L, cultivated 40 hours in the shaking table of the rotating speed under 200rpm and 30 ℃, the concentration that recorded cell with the dry cell weight method of masurement at the 10th hour is 87.21mg/L, uses phenol that liquid chromatograph measures and the concentration of meta-cresol to be respectively 281.15mg/L and 98.29mg/L.
Embodiment 9
The impact of meta-cresol concentration Degradation of Phenol
Dispose on request the minimal medium of phenols degraded, under 115 ℃, carry out the 30min sterilization.The fixing phenol that adds 600mg/L, and the meta-cresol of 100mg/L, cultivated 40 hours in the shaking table of the rotating speed under 200rpm and 30 ℃, the concentration that recorded cell with the dry cell weight method of masurement at the 15th hour is 163.26mg/L, uses phenol that liquid chromatograph measures and the concentration of meta-cresol to be respectively 12.11mg/L and 83.24mg/L.
The impact of meta-cresol concentration Degradation of Phenol
Dispose on request the minimal medium of phenols degraded, under 115 ℃, carry out the 30min sterilization.The fixing phenol that adds 600mg/L, and the meta-cresol of 100mg/L, cultivated 40 hours in the shaking table of the rotating speed under 200rpm and 30 ℃, the concentration that recorded cell with the dry cell weight method of masurement at the 20th hour is 364.41mg/L, uses phenol that liquid chromatograph measures and the concentration of meta-cresol to be respectively 0mg/L and 0mg/L.
Embodiment 11
The impact of meta-cresol concentration Degradation of Phenol
Dispose on request the minimal medium of phenols degraded, under 115 ℃, carry out the 30min sterilization.The fixing phenol that adds 600mg/L, and the meta-cresol of 500mg/L, cultivated 40 hours in the shaking table of the rotating speed under 200rpm and 30 ℃, the concentration that recorded cell with the dry cell weight method of masurement at the 10th hour is 45.48mg/L, uses phenol that liquid chromatograph measures and the concentration of meta-cresol to be respectively 598.14mg/L and 496.31mg/L.
Embodiment 12
The impact of meta-cresol concentration Degradation of Phenol
Dispose on request the minimal medium of phenols degraded, under 115 ℃, carry out the 30min sterilization.The fixing phenol that adds 600mg/L, and the meta-cresol of 500mg/L, cultivated 40 hours in the shaking table of the rotating speed under 200rpm and 30 ℃, the concentration that recorded cell with the dry cell weight method of masurement at the 20th hour is 170.63mg/L, uses phenol that liquid chromatograph measures and the concentration of meta-cresol to be respectively 312.54mg/L and 306.51mg/L.
Embodiment 13
The impact of meta-cresol concentration Degradation of Phenol
Dispose on request the minimal medium of phenols degraded, under 115 ℃, carry out the 30min sterilization.The fixing phenol that adds 600mg/L, and the meta-cresol of 500mg/L, cultivated 40 hours in the shaking table of the rotating speed under 200rpm and 30 ℃, the concentration that recorded cell with the dry cell weight method of masurement at the 30th hour is 329.15mg/L, uses phenol that liquid chromatograph measures and the concentration of meta-cresol to be respectively 0mg/L and 129.46mg/L.
Test data is drawn such as Fig. 1 and Fig. 2: can clearly draw from figure: the interaction in the polyphenol pollutent biodegradation process, utilize in the process of Pseudomonas aeruginosa GIMT1.074 degradation of phenol and meta-cresol mixture and find:
1. exist mutual inhibition and promotion effect between two kinds of substrates, the restraining effect of the higher Degradation of Phenol of concentration of meta-cresol is larger, and it is longer that cell concn reaches the used time of same OD value, but after the reaction times of abundance, meta-cresol concentration is higher, and final cell concentration is larger; Simultaneously, the concentration of meta-cresol is higher, and the needed time of degradable phenol and meta-cresol is longer.
2. the concentration of phenol is higher, and degraded has certain restraining effect to meta-cresol, and it is shorter that cell concn reaches the used time of same OD value, but after the reaction times of abundance, phenol concentration is higher, and final cell concentration is larger; Simultaneously, the concentration of phenol is higher, and the degradation time of degradable meta-cresol and phenol is longer.
Claims (8)
1. the interactional research method in the polyphenol pollutent biodegradation process is characterized in that step is as follows:
1) configuration is used for the substratum of degraded polyphenol pollutent; The used selective medium of phenols degradation experiment is to add phenol or meta-cresol on the minimal medium basis as carbon source; At the different meta-cresol concentration Degradation of Phenols of research when affect, the fixing concentration of phenol, several gradient levels of the concentration different concns of selection meta-cresol; When the different phenol concentration of research affects the degraded of meta-cresol, the fixing concentration of meta-cresol, several gradient levels of the different concns of selection phenol; Each gradient is carried out parallel laboratory test;
2) utilizing Pseudomonas aeruginosa GIMT1.074 to be inoculated in the substratum cultivates;
3) adopt optical densitometric method and constant weight desiccating method to measure cell concentration; Adopt the aldehydes matter concentration in the high effective liquid chromatography for measuring degraded system;
4) according to the variation of cell concn in the culturing process and the variation of phenol and meta-cresol degraded situation, obtain the interaction data in the polyphenol pollutent biodegradation process.
2. the method for claim 1 is characterized in that described substratum is: 500mg (NH
4)
2SO
4, 200mgKH
2PO
4, 100mg MgSO
4, 800mg Na
2HPO
412H
2O, the 20mg yeast soaks powder, 10mL trace elements mother liquor and 1000mL H
2O.
3. method as claimed in claim 2 is characterized in that described trace elements mother liquor composition is as follows: 0.4gMnSO
44H
2O, 0.4g ZnSO
47H
2O, 0.1gNa
2MoO
42H
2O, 0.1g CuSO
45H
2O, 1.0g CaCl
2, 10.0g Na
2SO
4, 2.0g FeSO
47H
2O, 0.5mL H
2SO
4, 2.0g NaOH, 12.0g disodium ethylene diamine tetraacetate and 1000mL H
2O.
4. the method for claim 1 is characterized in that step 2) cultural method be: with pure initial OD
600Value is that 0.01 Pseudomonas aeruginosa GIMT1.074 is inoculated in the substratum, and all initial pH value of medium are 7.2, and liquid culture and solid culture temperature are 30 ℃, and rotating speed was 200rpm when shaking table was cultivated, and incubation time is 40 hours.
5. the method for claim 1, it is characterized in that optical densitometric method measures the cell concentration method and be: optical density(OD) is done blank with corresponding substratum, measures the absorbancy (OD of bacteria suspension under the 600nm wavelength
600); The concentration of sample need to be diluted to multiply each other with measured result and extension rate below 0.8 when surpassing 0.8 again and obtain the actual OD of bacterium liquid
600Value.
6. the method for claim 1 is characterized in that constant weight dry method measurement cell concentration method is: to dry weight, the bacterium liquid of certain volume physiological period different concns got in the weighing record with resistant to elevated temperatures centrifuge tube constant weight, and centrifugal, supernatant liquor inclines; Wash cell with sterile distilled water, repetitive scrubbing twice is abandoned supernatant, then will be dried to constant weight, the weighing record in centrifuge tube and the thalline placement loft drier.
7. the method for claim 1, it is characterized in that high performance liquid chromatography is: high performance liquid chromatography chromatogram model is Agilent high performance liquid chromatography 1200, the chromatographic column model is Eclipse XDB-C18,4.6 * 250mm, 5 μ m; Moving phase adopts volume ratio methyl alcohol: water=4:3, flow velocity 1.0mL/min; Sample size is 10 μ L, 30 ℃ of column temperatures, and the UV device detects wavelength 280nm.
8. the interaction in the polyphenol pollutent biodegradation process is characterized in that utilizing in the process of Pseudomonas aeruginosa GIMT1.074 degradation of phenol and meta-cresol mixture and finds:
1) restraining effect of the higher Degradation of Phenol of concentration of meta-cresol is larger, and it is longer that cell concn reaches the used time of same OD value, but after the reaction times of abundance, meta-cresol concentration is higher, and final cell concentration is larger; Simultaneously, the concentration of meta-cresol is higher, and the needed time of degradable phenol and meta-cresol is longer;
2) concentration of phenol is higher, and degraded has certain restraining effect to meta-cresol, and it is shorter that cell concn reaches the used time of same OD value, but after the reaction times of abundance, phenol concentration is higher, and final cell concentration is larger; Simultaneously, the concentration of phenol is higher, and the degradation time of degradable meta-cresol and phenol is longer.
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| CN103773727A (en) * | 2014-02-11 | 2014-05-07 | 河南中烟工业有限责任公司 | Application of sphingomonas strain xp in degrading polyphenolic compounds in tobacco stems |
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