CN101974433A - Carbendazol composite degrading bacteria and method for degrading carbendazol - Google Patents
Carbendazol composite degrading bacteria and method for degrading carbendazol Download PDFInfo
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- CN101974433A CN101974433A CN 201010513256 CN201010513256A CN101974433A CN 101974433 A CN101974433 A CN 101974433A CN 201010513256 CN201010513256 CN 201010513256 CN 201010513256 A CN201010513256 A CN 201010513256A CN 101974433 A CN101974433 A CN 101974433A
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- derosal
- carbendazol
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Abstract
The invention discloses carbendazol composite degrading bacteria and a method for degrading carbendazol. The carbendazol composite degrading bacteria consist of alcaligenes M-1 and rhodococcus erythropolis L-1 in a compound ratio of 1:4. In the method for degrading the carbendazol, the inoculum concentration is 7 percent; the pH of the degrading condition is 6; the temperature is 30 DEG C; and the degrading time is 24 hours. The degrading rate of the carbendazol is up to 75.76 percent in 24 hours.
Description
Technical field
The present invention relates to the method for a kind of derosal composite degradation bacterium and degraded derosal, belong to biological high-technology field, be to utilize method of microorganism degraded chemical pesticide residual, be applicable to the production of the biological restoration of derosal contaminated soil and the green non-pollution agricultural-food in the modern agriculture.
Background technology
In agriculture production, people are extensive use of organic class agricultural chemicals such as various sterilants, sterilant, miticide and weedicide.Wherein, derosal is that demand is the sterilant kind of ton, is the principal item of China's agricultural chemicals outlet.From 1967 by since the du pont company development, derosal is widely used in the fungus-proof antisepsis by fungus-caused crop pest and Industrial materials.But, the derosal chemical property is stable, degradation half life is longer in physical environment, can produce certain restraining effect to the richness and the dominance of microflora in the soil, and its residual build-up effect by food chain in fruit, plant, soil influences human health.
Because a large amount of for many years derosal that use continuously make many phytopathogens produce resistance to it.In order to reach prevention effect, the peasant has strengthened the consumption of derosal in use, has caused the problem of its residual quantity severe overweight in agricultural-food, becomes the barrier of the green agricultural byproducts export trade of China.2007, EU Committee was adjusted into 0.05mg/kg with the maximum residue limit (MRL) of derosal in oranges and tangerines by 5mg/kg; MRL in edible fungi is adjusted into 0.1mg/kg by 1mg/kg.
At present, the application in agriculture of chemical pesticide is an alternate fully, therefore, how effectively to remove the pollution problem that chemical pesticide brings and is subjected to common concern.Handle with respect to physical chemistry, it is a kind of Pollution abatement approach more effective, that more save that microorganism is repaired pesticide residue, also can bring significant social and ecological benefits.
Summary of the invention
Technical problem to be solved by this invention is to provide a kind of derosal composite degradation bacterium and a kind of method of using this composite degradation bacterium degraded derosal at the deficiencies in the prior art.
Derosal composite degradation bacterium provided by the invention is made up of two kinds of bacterium, is respectively Alcaligenes M-1 (Alcaligenes sp.), is numbered A and rhodococcus erythropolis L-1 (Rhodococcus erythropolis), is numbered R.
Alcaligenes M-1 (Alcaligenes sp.) separates from the Yang Ling area of long-term application derosal plastic shed soil.This bacterial strain can be sole carbon source growth with the derosal, and bacterium colony is rounded, oyster white, neat in edge, surface elevation, moistening smooth.Thalline is rod-short, Gram-negative, methyl red and V.P. reaction negative.Bacterium colony on agar is chromogenesis not, oxydase, the catalase positive.Edwardsiella hoshinae can not hydrolyzed starch, can gelatin hydrolysate, and the nitrate reduction experiment is positive.Tentatively this bacterial strain is defined as Alcaligenes (Alcaligenes sp.) through 16S rDNA sequential analysis again.The degraded optimum pH is 6.0, and optimum temperuture is 25 ℃, can degrade the derosal of 200mg/L more than 90% in 72 hours.
Rhodococcus erythropolis L-1 (Rhodococcus erythropolis) separates the former pharmaceutical factory sewage sludge of producing derosal certainly for many years.But this bacterium also derosal is sole carbon source and energy growth, and bacterium colony takes on a red color, opaque circle, and is smooth, surface elevation, and moistening, the edge is complete.The degraded optimum pH is 6.0, and optimum temperuture is 30 ℃, and the starting point concentration of can degrading fully was the derosal of 200mg/L in 72 hours.Its physio-biochemical characteristics are as follows:
The physio-biochemical characteristics of table 1 bacterial strain L-1
+：POSITIVE；-：NEGATIVE
Embodiment
Below in conjunction with specific embodiment, the present invention is described in detail.
Substratum: 1. LB liquid nutrient medium: NaCl 10.0g, peptone 10.0g, yeast soak powder 5.0g, distilled water 1000mL, pH 7.0.2. minimal medium: NaCl 1.0g, K
2HPO
41.5g,, KH
2PO
40.5g, (NH
4)
2SO
42.0g, MgSO
47H
2O 0.2g, distilled water 1000mL, pH 7.0.
Reagent and instrument: derosal (Petroleum Chemical Engineering Institute provides by Shaanxi, purity 98%); UV-1200 type ultraviolet-visible pectrophotometer reaches Instr Ltd. available from the U.S. spectrum in Shanghai; SKY-200B type constant temperature culture vibrator is available from Sukun Industry Co., Ltd., Shanghai; High performance liquid chromatograph LC-2010AHT is available from day island proper Tianjin company.
Derosal Determination on content method:
The three-wavelength correction method: the derosal standardized solution of preparation 5g/L (effective constituent) is diluted to respective concentration as required.Pipette 100 μ L samples, (1+11) is settled to 10mL with dilute hydrochloric acid.Measure it respectively 278,281 and the absorbancy at 290nm wavelength place, calculate by Δ A=A281-(A278+A290)/2 and proofread and correct absorbancy Δ A, the derosal content in the sample is carried out quantitative analysis according to typical curve.
High performance liquid chromatography: referring to the agricultural industry criteria NY/T 1680-2009 of the People's Republic of China (PRC).
The preparation of seed liquor:
Two kinds of bacterial strains that are stored in the inclined-plane are activated and tame cultivation respectively, and picking one ring thalline is seeded in the LB liquid nutrient medium, and 30 ℃ shake cultivation 24h down.Adjust its OD600 before the use and be 1.0, pipette 5mL bacterium liquid, at 4 ℃, centrifugal 5min under the 4000r/min collects thalline, after the inorganic salt liquid substratum washing of sterilizing 1 time, suspends with isopyknic sterilization inorganic salt liquid substratum, and is standby.
Single bacterium and composite bacteria are tested the degraded of derosal:
Add the derosal standardized solution in the inorganic salt liquid substratum, making its concentration is 200mg/L, totally is 100mL, and regulating pH is 6.0.Inoculum size by 5% inserts single bacterium A, R and composite bacteria AR (compound proportion is 1: 4), is blank to insert sterilized water, each processing be provided with 3 parallel.Place 30 ℃, shaking table shaking culture under the 150r/min.The result is as shown in table 2:
Single bacterium of table 2 and composite bacteria are to the degraded of derosal
Composite bacteria is to the degraded of different concns derosal
In minimal medium, add the derosal standardized solution, make that its concentration is respectively 100,200,300,400,500,600mg/L, regulating pH value is 6.0, and the inoculum size with 5% inserts composite flora AR (compound proportion is 1: 4), shaking table cultivation under 30 ℃, the condition of 150r/min.The result is as shown in table 3:
Table 3 composite bacteria is to the degraded of different concns derosal
The test of composite bacteria orthogonal optimization
This experiment is investigated the influence to composite bacteria degraded derosal of temperature, pH value and three factors of inoculum size on the basis of single bacterium to the degradation characteristic research of derosal.Adopt three factors, three horizontal quadrature experimental programs, according to L
9(3
4) orthogonal table, the row unrepeated test of having time.Utilization intuitive analysis method and analysis of variance are assessed the influence degree of each factor, and provide optimum combination condition.Best of breed is the 5th group of test, and promptly temperature is 30 ℃, and inoculum size is 7%, and pH6.0, composite bacteria AR are AR (1: 4)
Measurement result is as follows:
Single bacterium of table 4 and composite bacteria are to the comparison of different concns derosal degradation rate
Table 5 orthogonal experiment data is handled
Table 6 orthogonal test The results of analysis of variance
| Source? | df? | SS? | MS? | F? | Pr＞F? |
| Temperature | 2? | 22.2998? | 11.1499? | 0.43? | 0.7005? |
| pH? | 2? | 2924.04816? | 1462.0240? | 56.08? | 0.0175? |
| Inoculum size | 2? | 23.9013? | 11.9506? | 0.46? | 0.6857? |
Should be understood that, for those of ordinary skills, can be improved according to the above description or conversion, and all these improvement and conversion all should belong to the protection domain of claims of the present invention.
Claims (4)
1. a derosal composite degradation bacterium is characterized in that, this composite degradation bacterium is made up of two kinds of bacterium, is respectively Alcaligenes M-1 and rhodococcus erythropolis L-1.
2. derosal composite degradation according to claim 1 is characterized in that, the compound proportion of described Alcaligenes M-1 and described rhodococcus erythropolis L-1 is 1: 4.
3. method of using the described derosal composite degradation of claim 1 bacterium degraded derosal.
4. the method for degraded derosal according to claim 3 is characterized in that, inoculum size 7%, and degradation condition is PH=6, and temperature is 30 ℃, and degradation time is 24h.
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102296041A (en) * | 2011-08-26 | 2011-12-28 | 中国农业科学院农业资源与农业区划研究所 | Bacterium for efficiently degrading residual pesticide carbendazim and application thereof |
| CN104862245A (en) * | 2015-04-16 | 2015-08-26 | 西北农林科技大学 | Carbendazim degrading bacteria MBC-6F and application thereof |
| CN107446855A (en) * | 2017-08-29 | 2017-12-08 | 西安罗格斯生物科技有限公司 | The application of Alcaligenes bacterial strain, enzyme preparation and its degrading pesticide residues |
| CN115739973A (en) * | 2022-11-29 | 2023-03-07 | 湖南农业大学 | Method for treating carbendazim contaminated soil by combining mushroom dregs with nitrification inhibitor |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101260378A (en) * | 2008-04-28 | 2008-09-10 | 上海交通大学 | Application of rhodococcus erythropolis in cleaning heavy oil dirt |
| CN101701222A (en) * | 2009-04-17 | 2010-05-05 | 华东理工大学 | Gene for coding nitrilase of alcaligenes and method for preparing single enantiomorph of mandelic acid using same |
-
2010
- 2010-10-21 CN CN 201010513256 patent/CN101974433A/en active Pending
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101260378A (en) * | 2008-04-28 | 2008-09-10 | 上海交通大学 | Application of rhodococcus erythropolis in cleaning heavy oil dirt |
| CN101701222A (en) * | 2009-04-17 | 2010-05-05 | 华东理工大学 | Gene for coding nitrilase of alcaligenes and method for preparing single enantiomorph of mandelic acid using same |
Non-Patent Citations (1)
| Title |
|---|
| 《华中农业大学学报》 20090531 张滨; 马美湖; 杨华 红平红球菌产微生物絮凝剂的发酵条件及分离纯化 全文 1－4 , * |
Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102296041A (en) * | 2011-08-26 | 2011-12-28 | 中国农业科学院农业资源与农业区划研究所 | Bacterium for efficiently degrading residual pesticide carbendazim and application thereof |
| CN102296041B (en) * | 2011-08-26 | 2013-07-31 | 中国农业科学院农业资源与农业区划研究所 | Bacterium for efficiently degrading residual pesticide carbendazim and application thereof |
| CN104862245A (en) * | 2015-04-16 | 2015-08-26 | 西北农林科技大学 | Carbendazim degrading bacteria MBC-6F and application thereof |
| CN104862245B (en) * | 2015-04-16 | 2017-09-19 | 西北农林科技大学 | One plant of carbendazim degrading bacterium MBC 6F and its application |
| CN107446855A (en) * | 2017-08-29 | 2017-12-08 | 西安罗格斯生物科技有限公司 | The application of Alcaligenes bacterial strain, enzyme preparation and its degrading pesticide residues |
| CN115739973A (en) * | 2022-11-29 | 2023-03-07 | 湖南农业大学 | Method for treating carbendazim contaminated soil by combining mushroom dregs with nitrification inhibitor |
| CN115739973B (en) * | 2022-11-29 | 2024-04-12 | 湖南农业大学 | Method for treating carbendazim-polluted soil by combining fungus dreg with nitrification inhibitor |
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Application publication date: 20110216 |