CN106277362A - Method with pycnoporus samguineus degraded sulfonamides compound - Google Patents
Method with pycnoporus samguineus degraded sulfonamides compound Download PDFInfo
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- CN106277362A CN106277362A CN201610780098.6A CN201610780098A CN106277362A CN 106277362 A CN106277362 A CN 106277362A CN 201610780098 A CN201610780098 A CN 201610780098A CN 106277362 A CN106277362 A CN 106277362A
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- pycnoporus
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- sulfonamides compound
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- C—CHEMISTRY; METALLURGY
- C02—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F3/00—Biological treatment of water, waste water, or sewage
- C02F3/34—Biological treatment of water, waste water, or sewage characterised by the microorganisms used
- C02F3/342—Biological treatment of water, waste water, or sewage characterised by the microorganisms used characterised by the enzymes used
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- C—CHEMISTRY; METALLURGY
- C02—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F2101/00—Nature of the contaminant
- C02F2101/30—Organic compounds
- C02F2101/38—Organic compounds containing nitrogen
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- C—CHEMISTRY; METALLURGY
- C02—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F2101/00—Nature of the contaminant
- C02F2101/30—Organic compounds
- C02F2101/40—Organic compounds containing sulfur
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Abstract
The invention discloses the method with pycnoporus samguineus degraded sulfonamides compound, comprise the steps: that the pycnoporus samguineus that deposit number is CGMCC 5.815 is cultivated by (1), obtain pycnoporus samguineus first order seed；(2) first order seed of pycnoporus samguineus is taken, centrifugal, abandon supernatant, with fermentation medium, thalline is resuspended, fermentation；(3) sulfonamides compound is produced waste discharge to mix with fermentation liquid, add 2,2 azino two (3 ethyl-benzothiazole 6 sulfonic acid) di-ammonium salts, degraded.The method of the present invention not only provides a kind of condition of culture that pycnoporus samguineus can be made quickly to produce laccase in the presence of not having inducer, and establish a kind of method utilizing whole thalline and extracellular environment thereof to carry out sulfonamides compound degraded in fermentation liquid, eliminate a series of loaded down with trivial details steps such as isolated and purified.Fermentation time is short, and degradation rate is high.Degradation process can be repeated exactly, sufficiently stable.
Description
Technical field
The invention belongs to field of environment pollution control, relate to the side of a kind of pycnoporus samguineus degraded sulfonamides compound
Method.
Background technology
Laccase (Laccases) is the protein of the multiple copper ion of a kind of combination, belongs to covellite oxidase.Laccase can be survived
In air, after reacting, unique product is exactly water, is the most substantially a kind of environment-friendly type ferment.Due to this environmental protection in several years
Consciousness is gradually paid attention to by people, and laccase also becomes the object of study of numerous scholar the most in recent years.Laccase is common in fungus and plants
In thing.Wherein, the laccase that whiterot fungi produces serves important function during lignin degradation, is particularly subject to pay close attention to.And,
The laccase that whiterot fungi produces mostly is exoenzyme, it is easy to extraction and application.The whiterot fungi of most study is Trametes Trametes at present
With pleurotus Pleurotus.In recent years, samguineus belongs to Pycnoporus to start to cause concern, because it can produce has height
The laccase of oxidation-reduction potential, it means that it is bigger that samguineus belongs to the substrate spectrum that can act on of laccase that Pycnoporus produces,
And laccase is samguineus belongs to the mainization ferment that Pycnoporus produces when liquid fermentation.
The technology of existing production laccase, needs longer fermentation period more, and especially at the fermentation initial stage, within 60h, laccase produces
Measure extremely low.Using natural medium to ferment, production process repeatability is poor.It addition, prior art utilizes laccase degraded sulphur
Aminated compounds, uses from thalline broth extraction purification or the laccase that slightly carries more, and this series of programs is loaded down with trivial details and of a high price.
Summary of the invention
It is an object of the invention to overcome the deficiencies in the prior art, it is provided that one pycnoporus samguineus is degraded sulfonamides compound
Method.
Technical scheme is summarized as follows:
By the method for pycnoporus samguineus degraded sulfonamides compound, comprise the steps:
(1) seed culture: by the pycnoporus samguineus (Pycnoporus that deposit number is CGMCC 5.815
Sanguineus) in potato dextrose agar solid medium, cultivate 6-8 days at 25-30 DEG C；Proceed to Rhizoma Solani tuber osi Fructus Vitis viniferae again
In sugar liquors culture medium, cultivate 45-50 hour at 25-30 DEG C, obtain pycnoporus samguineus first order seed；
(2) fermentation culture: take the first order seed of the pycnoporus samguineus that step (1) obtains, be centrifuged 8-at 6000-10000rpm
12min, abandons supernatant, with fermentation medium, thalline is resuspended, and at 25-30 DEG C, under the conditions of 100-150rpm, ferment 40-50h；Institute
The volume ratio stating first order seed and fermentation medium is 1:10-12.5；
(3) degraded sulfonamides compound: sulfonamides compound is produced waste discharge mixed with the fermentation liquid that step (2) obtains
Closing, the concentration making sulfonamides compound is 20-50mg/L, adds 2,2-azino-two (3-ethyl-benzothiazole-6-sulfonic acid) two
Ammonium salt, making concentration is 0.4-0.6mM, and degrade 5-8h.
Potato dextrose agar solid medium is preferably: Rhizoma Solani tuber osi 200g/L, glucose 20g/L, agar 18g/L,
Natural pH, surplus is water.
Potato dextrose broth is preferably: Rhizoma Solani tuber osi 200g/L, glucose 20g/L, and natural pH, surplus is
Water.
Preferably, fermentation medium is made by following method: take glucose, 1.0g/L；KH2PO4,1.0g/L；NaH2PO4,
0.13g/L；2,2-dimethyl succinate 1.0g/L；MgSO4·7H2O,0.5g/L；CuSO4·5H2O,0.01g/L；CaCl2·
2H2O,0.05g/L；ZnSO4·7H2O,0.03g/L；MnSO4·H2O,0.02g/L；CoCl2·6H2O,0.001g/L；Sodium acetate
1g/L, is dissolved in water, sterilizing, obtains solution one；Take (NH4)2SO4,0.5g/L；FeSO4·7H2O,0.01g/L；It is dissolved in water, makes
With the membrane filtration that aperture is 0.22 μm, obtain solution two；Solution one and solution two are mixed, fills up excess water.
Sulfonamides compound is preferably sulfamethoxazole, sulfadiazine, bacteresulf, sulfamethazine or sulfanilamide
SDM.
Advantages of the present invention:
The method of the present invention not only provides one can make P.sanguineus in the presence of not having inducer
Quickly produce the condition of culture of laccase, and establish one and utilize whole thalline and extracellular environment thereof to carry out sulfanilamide in fermentation liquid
The method of compounds degraded, eliminates a series of loaded down with trivial details steps such as isolated and purified.Fermentation time of the present invention is greatly shortened, fall
Solution rate also significantly improves.Further, in the conditions of the invention, degradation process can be repeated exactly, sufficiently stable.
Accompanying drawing explanation
Fig. 1 is the situation that pycnoporus samguineus produces laccase during the fermentation.
Fig. 2 is the situation of pycnoporus samguineus degraded sulfamethoxazole (SMX).
Detailed description of the invention
2,2-azino-two (3-ethyl-benzothiazole-6-sulfonic acid) di-ammonium salts English is, 2-azino-bis-(3-
ethylbenzothiazoline-6-sulfonic acid)(ABTS)
Below in conjunction with specific embodiment, the present invention is further described:
In each embodiment:
Potato dextrose agar solid medium is: Rhizoma Solani tuber osi 200g/L, glucose 20g/L, agar 18g/L, natural
PH, surplus is water.
Potato dextrose broth is: Rhizoma Solani tuber osi 200g/L, glucose 20g/L, and natural pH, surplus is water.
Fermentation medium is made by following method: take glucose, 1.0g/L；KH2PO4,1.0g/L；NaH2PO4,0.13g/
L；2,2-dimethyl succinate 1.0g/L；MgSO4·7H2O,0.5g/L；CuSO4·5H2O,0.01g/L；CaCl2·2H2O,
0.05g/L；ZnSO4·7H2O,0.03g/L；MnSO4·H2O,0.02g/L；CoCl2·6H2O,0.001g/L；Sodium acetate 1g/L,
It is dissolved in water, sterilizing, obtain solution one；Take (NH4)2SO4,0.5g/L；FeSO4·7H2O,0.01g/L；It is dissolved in water, uses aperture
It is the membrane filtration of 0.22 μm, obtains solution two；Solution one and solution two are mixed, fills up excess water.
Embodiment 1
By the method for pycnoporus samguineus degraded sulfonamides compound, comprise the steps:
(1) seed culture: by the pycnoporus samguineus (Pycnoporus that deposit number is CGMCC 5.815
Sanguineus) in potato dextrose agar solid medium, cultivate 7 days at 28 DEG C；Proceed to potato glucose liquid again
In body culture medium, cultivate 48 hours at 28 DEG C, obtain pycnoporus samguineus first order seed；
(2) fermentation culture: take the first order seed 40mL of the pycnoporus samguineus that step (1) obtains, is centrifuged at 8000rpm
10min, abandons supernatant, with 460mL fermentation medium, thalline is resuspended, and at 28 DEG C, under the conditions of 130rpm, ferment 48h, laccase activity
For 0.85U/mL, see Fig. 1；
(3) degraded sulfonamides compound: sulfamethoxazole is produced waste discharge mixed with the fermentation liquid that step (2) obtains
Closing, the concentration making sulfamethoxazole is 50mg/L, addition 2,2-azino-two (3-ethyl-benzothiazole-6-sulfonic acid) di-ammonium salts,
Making concentration is 0.5mM, and degrade 6h.(see Fig. 2,54 hours in Fig. 2 are 48h and the 6h sums of step (3) of step (2)) sulfanilamide
First oxazole (SMX) degradation rate is 100%.
Embodiment 2
By the method for pycnoporus samguineus degraded sulfonamides compound, comprise the steps:
(1) seed culture: by the pycnoporus samguineus (Pycnoporus that deposit number is CGMCC 5.815
Sanguineus) in potato dextrose agar solid medium, cultivate 8 days at 25 DEG C；Proceed to potato glucose liquid again
Body culture medium is cultivated 50 hours at 25 DEG C, obtains pycnoporus samguineus first order seed；
(2) fermentation culture: take the first order seed 45mL of the pycnoporus samguineus that step (1) obtains, is centrifuged at 6000rpm
12min, abandons supernatant, with 450mL fermentation medium, thalline is resuspended, and at 25 DEG C, under the conditions of 100rpm, ferment 50h；Laccase activity
For 0.78U/mL；
(3) degraded sulfonamides compound: sulfadiazine is produced the fermentation liquid that waste discharge obtains with step (2) and mixes,
The concentration making sulfadiazine is 20mg/L, addition 2, and 2-azino-two (3-ethyl-benzothiazole-6-sulfonic acid) di-ammonium salts makes concentration
For 0.4mM, degrade 5h.
Sulfadiazine degradation rate is 100%.
Embodiment 3
By the method for pycnoporus samguineus degraded sulfonamides compound, comprise the steps:
(1) seed culture: by the pycnoporus samguineus (Pycnoporus that deposit number is CGMCC 5.815
Sanguineus) in potato dextrose agar solid medium, cultivate 6 days at 30 DEG C；Proceed to potato glucose liquid again
Body culture medium is cultivated 45 hours at 30 DEG C, obtains pycnoporus samguineus first order seed；
(2) fermentation culture: take the first order seed 40mL of the pycnoporus samguineus that step (1) obtains, is centrifuged at 10000rpm
8min, abandons supernatant, with 500mL fermentation medium, thalline is resuspended, and at 30 DEG C, under the conditions of 150rpm, ferment 40h；Laccase activity
For 0.92U/mL；
(3) degraded sulfonamides compound: bacteresulf is produced waste discharge mixed with the fermentation liquid that step (2) obtains
Closing, the concentration making bacteresulf is 35mg/L, addition 2,2-azino-two (3-ethyl-benzothiazole-6-sulfonic acid) di-ammonium salts,
Making concentration is 0.6mM, and degrade 8h.
Bacteresulf degradation rate is 100%.
It is demonstrated experimentally that substitute the bacteresulf of the present embodiment with sulfamethazine or sulfadimethoxine, its
Its same the present embodiment, its degradation rate is 100%.
Chromatographic column used by high performance liquid chromatography is C18 chromatographic column, and instrument is Waters e2695Alliance HPLC
With 2489UV/Visible detector, detection wavelength is 265nm.Testing conditions is: chromatogram column temperature 30 DEG C, sampling volume 10 μ
L, flow velocity 1mL min-1.Flowing phase composition: 40% acetonitrile and 60%0.1% formic acid solution.
Claims (5)
1., by the method for pycnoporus samguineus degraded sulfonamides compound, its feature comprises the steps:
(1) seed culture: the pycnoporus samguineus (Pycnoporus sanguineus) that deposit number is CGMCC 5.815 is existed
In potato dextrose agar solid medium, cultivate 6-8 days at 25-30 DEG C；Proceed to potato dextrose broth again
In, cultivate 45-50 hour at 25-30 DEG C, obtain pycnoporus samguineus first order seed；
(2) fermentation culture: take the first order seed of the pycnoporus samguineus that step (1) obtains, be centrifuged 8-at 6000-10000rpm
12min, abandons supernatant, with fermentation medium, thalline is resuspended, and at 25-30 DEG C, under the conditions of 100-150rpm, ferment 40-50h；Institute
The volume ratio stating first order seed and fermentation medium is 1:10-12.5；
(3) degraded sulfonamides compound: sulfonamides compound is produced the fermentation liquid that waste discharge obtains with step (2) and mixes,
The concentration making sulfonamides compound is 20-50mg/L, adds 2,2-azino-two (3-ethyl-benzothiazole-6-sulfonic acid) diammonium
Salt, making concentration is 0.4-0.6mM, and degrade 5-8h.
Method the most according to claim 1, is characterized in that described potato dextrose agar solid medium is: Rhizoma Solani tuber osi
200g/L, glucose 20g/L, agar 18g/L, natural pH, surplus is water.
Method the most according to claim 1, is characterized in that described potato dextrose broth is: Rhizoma Solani tuber osi
200g/L, glucose 20g/L, natural pH, surplus is water.
Method the most according to claim 1, is characterized in that described fermentation medium is made by following method: take glucose,
1.0g/L；KH2PO4,1.0g/L；NaH2PO4,0.13g/L；2,2-dimethyl succinate 1.0g/L；MgSO4·7H2O,0.5g/L；
CuSO4·5H2O,0.01g/L；CaCl2·2H2O,0.05g/L；ZnSO4·7H2O,0.03g/L；MnSO4·H2O,0.02g/L；
CoCl2·6H2O,0.001g/L；Sodium acetate 1g/L, is dissolved in water, sterilizing, obtains solution one；Take (NH4)2SO4,0.5g/L；
FeSO4·7H2O,0.01g/L；It is dissolved in water, the membrane filtration using aperture to be 0.22 μm, obtain solution two；By solution one and molten
Liquid two mixes, and fills up excess water.
Method the most according to claim 1, is characterized in that described sulfonamides compound is that sulfamethoxazole, sulfanilamide are phonetic
Pyridine, bacteresulf, sulfamethazine or sulfadimethoxine.
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108217973A (en) * | 2018-01-04 | 2018-06-29 | 南方科技大学 | A kind of efficient degradation method of sulphadiazine |
| CN110526418A (en) * | 2019-08-19 | 2019-12-03 | 江南大学 | A method of utilizing antibiotic in immobilization samguineus degrading cultivation waste water |
| CN116425316A (en) * | 2023-03-10 | 2023-07-14 | 中国水产科学研究院黄海水产研究所 | Application of sperm whale Mb and mutant H64D/V68I Mb in degradation of sulfonamide compounds |
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| CN1621519A (en) * | 2003-11-25 | 2005-06-01 | 中国科学院微生物研究所 | Method for preparing pycnoporus samguineus GW fungal laccase |
| CN104561105A (en) * | 2014-12-15 | 2015-04-29 | 华南理工大学 | Method for synthesizing gold nanoparticles by using pycnoporus sanguineu organisms and application of method for synthesizing gold nanoparticles by using pycnoporus sanguineu organism |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN1621519A (en) * | 2003-11-25 | 2005-06-01 | 中国科学院微生物研究所 | Method for preparing pycnoporus samguineus GW fungal laccase |
| CN104561105A (en) * | 2014-12-15 | 2015-04-29 | 华南理工大学 | Method for synthesizing gold nanoparticles by using pycnoporus sanguineu organisms and application of method for synthesizing gold nanoparticles by using pycnoporus sanguineu organism |
Non-Patent Citations (1)
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| XIN LI等: "Improving the bioremoval of sulfamethoxazole and alleviating cytotoxicity of its biotransformation by laccase producing system under coculture of Pycnoporus sanguineus and Alcaligenes faecalis", 《BIORESOURCE TECHNOLOGY》 * |
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108217973A (en) * | 2018-01-04 | 2018-06-29 | 南方科技大学 | A kind of efficient degradation method of sulphadiazine |
| CN110526418A (en) * | 2019-08-19 | 2019-12-03 | 江南大学 | A method of utilizing antibiotic in immobilization samguineus degrading cultivation waste water |
| CN116425316A (en) * | 2023-03-10 | 2023-07-14 | 中国水产科学研究院黄海水产研究所 | Application of sperm whale Mb and mutant H64D/V68I Mb in degradation of sulfonamide compounds |
| CN116425316B (en) * | 2023-03-10 | 2023-11-21 | 中国水产科学研究院黄海水产研究所 | Application of sperm whale Mb and mutant H64D/V68I Mb in degradation of sulfonamide compounds |
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