JPH0220234B2 - - Google Patents

Description

[Detailed description of the invention]

The present invention relates to a method for decomposing ethylenediaminetetraacetic acid by bacteria. Ethylenediaminetetraacetic acid (hereinafter abbreviated as EDTA) is known as a chelating agent that forms strong complex compounds with various metals.
It is used in large quantities in industries, concentrated water industries, medicine, and other fields, but if EDTA is contained in wastewater from factories, it can cause environmental pollution such as eutrophication of seawater and lake water. Therefore, an effective decomposition treatment for EDTA is desired, but EDTA has been known as a substance that is difficult to decompose by microorganisms. It is known that EDTA can be decomposed by chemical treatments such as ultraviolet light and active chlorine (for example, see Japanese Patent Application Laid-open No. 51-31052), but this method is difficult to control, requires specific equipment, and is expensive. It has drawbacks such as being relatively expensive. Also EDTA
microorganisms in the soil [see Teisier's Applied Macrobiology Vol. Microbiology Vol. 29 No. 6 No. 787-794 (1975)], activated sludge (Yonezawa et al. Pollution Resources Research Institute Vol. 6 No. 4 No. 15-21 (1975) )], but despite long-term cultivation and treatment for several weeks, its decomposition power is extremely weak and is not practical, and there is no effective method for decomposing EDTA using microorganisms. From this perspective, the present inventors have widely studied the natural world.
As a result of searching for EDTA-assimilating bacteria, it was found that there are bacteria belonging to the genus Pseudomonas and bacteria belonging to the genus Alcaligeness that can grow in a medium using EDTA as a sole carbon source and a sole nitrogen source, and by bacteria
The present invention was achieved by discovering that EDTA is assimilated and decomposed. That is, the present invention is characterized in that bacteria belonging to the genus Pseudomonas or bacteria belonging to the genus Alcaligenes, which have the ability to decompose EDTA, are brought into contact with EDTA, a metal complex thereof, or a salt thereof. Here, EDTA assimilation refers to the phenomenon in which bacterial cells take in EDTA as a carbon source and nitrogen source, consume it, and proliferate, and the carbon-carbon bond and carbon-nitrogen bond of EDTA are simply broken. It does not mean that. Bacteria used in the present invention can be collected from a wide range of sources in the natural world, such as soil, river water, and seawater, but specifically, for example, Pseudomonas No.51-Y
(Feikoken Bibori No. 6103), Alcaligenes No. 51-z (Feikoku Kenbibi No. 6104)
can be mentioned. The mycological properties of Pseudomonas No. 51-Y and Alcaligenes No. 51-z are as shown in Table 1. Mycological testing and classification is carried out by Bergey's Manual of Bacteriological Bacteriology.
Manual of Determinative Bactariology) No. 7
8th edition.

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[Table] Based on the above mycological properties, Pseudomonas No.
51-Y strain and Alcaligenes No. 51-z strain were identified. Pseudomonas No. 51-Y strain was identified by Verge Ace, Manual of Determinative Research, based on its morphology, microscopic examination such as Gram staining, various physiological properties, and the fact that it was isolated from seawater.
Based on Bacteriology 7th and 8th editions,
It was identified as a bacterium closely related to Pseudomonas marinoglutinosa. However, Pseudomonas No. 51
-Y strain is clearly distinguished from Pseudomonas marinoglutinosa in that it does not liquefy gelatin or decompose starch. In addition, Alcaligenes No. 51-Z strain was identified as a bacterium belonging to the genus Alcaligenes based on its morphology, microscopic examination such as Gram staining, and various physiological properties.
It was identified as a bacterium similar to Alcaligenes faecalis described in the classification of Verge Ace, Manual of Determinative Bacteriology, 7th and 8th editions. The present invention was made based on the above discovery.
The present invention provides a method for microbial decomposition of EDTA, which is characterized by bringing these bacteria into contact with EDTA. EDTA in the present invention refers to the free acid of EDTA, magnesium, calcium, iron, cobalt, zinc,
It includes complex compounds with copper, etc., and salts of sodium, potassium, ammonium, etc., and these compounds may be present alone or in a mixture in the object to be treated. Furthermore, the degree of decomposition of EDTA according to the present invention is
It is indicated by the degree of decrease in EDTA or the degree of proliferation of bacterial cells. The present invention uses, for example, EDTA as a carbon source and nitrogen source, and a medium containing inorganic salts such as phosphate, potassium, magnesium, calcium, iron, manganese, zinc, and molybdenum (EDTA-containing liquid to be treated).
This can be carried out by growing the bacteria in the medium. In this case, if necessary, organic micronutrients such as yeast extract may be added to promote bacterial growth. Phosphate and potassium ions are used in the conventional form of potassium monohydrogen phosphate or potassium dihydrogen phosphate. Inorganic metal salts such as calcium, magnesium, iron, manganese, zinc, and molybdenum are used in the form of chlorides, sulfates, nitrides, oxides, and the like. It is not preferable to inoculate the bacteria into a medium rich in nutrients other than EDTA, such as peptone or meat extract, as the ability to decompose EDTA may decrease. The appropriate culture or treatment temperature is 20 to 37°C, and the pH of the medium (EDTA-containing treatment liquid) is 5.0 to 9.0, but more preferably the culture temperature is 30°C, and the pH of the medium is 5.0 to 9.0.
Around 7.0 is good. Cultivation or treatment is carried out by standing still, shaking or aerating, and the rate of assimilation of EDTA is rapid under conditions where oxygen is highly consumed. Bacteria can be grown well even in the dark or in closed culture tanks, where there is no light, but under light conditions, the growth rate of the bacteria is even faster, and it is usually cultured within 4 to 7 days. The concentration of EDTA in the medium can be decreased over time. Furthermore, in carrying out the method of the present invention, a culture obtained by culturing these microorganisms in an appropriate medium in advance, or a cell fraction concentrated by centrifugation, filtration, etc., if necessary, may be brought into contact with EDTA. Able to achieve purpose. In this case, it is not necessary to grow the bacteria, but it is preferable to keep the composition of the solution uniform with appropriate stirring to decompose EDTA. The amount of EDTA described in the present invention is determined by gas chromatography (Ruding et al., Water Research Bergamon Press, Vol. 6, No. 871).
876, 1972), and the percentage of EDTA reduced was expressed as the decomposition rate. Bacterial counts were also measured using a microscope. Next, the present invention will be explained by giving examples. Example 1 Disodium EDTA dihydrate 1.86g Dipotassium phosphate (K ₂ HPO ₄ ) 8.7g Magnesium sulfate (MgSO ₄ 7H ₂ O) 0.24g Calcium chloride (CaCl ₂ 2H ₂ O) 0.5mg Manganese chloride ( Dissolve 3 mg of zinc nitrate (ZnSO _{4.7H 2} O ₎ in 1000 ml _of distilled water and PH using a caustic sword.
Adjusted to 7.0. 50 ml of this was dispensed into a 500 ml Sakaguchi flask and sterilized at 120°C for 110 minutes to obtain a culture solution. A loopful of Pseudomonas strain No. 51-Y (Feikokenbacterium No. 6103) was inoculated into this medium, and the flask was completely covered with a black cloth that did not allow light to pass through.
The cells were cultured on a reciprocating shaker for hours. Table 2 shows the results of measuring the decomposition rate of EDTA and the number of bacteria. As a control example, when the medium composition and culture conditions are the same and no bacteria are inoculated, EDTA
No decrease was observed at all. Example 2 Among the culture medium composition, culture method, and culture conditions described in Example 1, the inoculated bacteria were used as Alcaligenes No. 51-
Table 2 shows the results of measuring the reduction of EDTA and the number of bacteria in the same manner as in Example 1 except that the strain was replaced with Z strain (Feikoken Bacteria No. 6104).

[Table] Example 3 Of the medium composition, culture method, and culture conditions described in Example 1, the EDTA2 sodium salt used in the medium was EDTA2 ammonium salt. In addition, when culturing with light, the culture was carried out in the same manner as in Example 1 except that the Sakaguchi flask was not covered with a black cloth and the culture was irradiated with fluorescent light at 600 to 800 lux. As a result of measuring the reduction and number of bacteria, it was found that culturing with light exposure had a significant effect on the decomposition rate of EDTA and the number of bacteria in the culture solution compared to conditions without light exposure. Table 3 shows the decomposition rate of EDTA and the change over time in the number of bacteria in the culture solution.

[Table] Example 4 EDTA disodium dihydrate 18.6g K ₂ HPO ₄ 87.0g MgSO ₄・7H ₂ O 2.4g Ca(NO ₃ ) ₂・4H ₂ O 5mg MnCl ₂・4H ₂ O 30mg ZnSO ₄・7H Dissolve 100mg of _2O in distilled water and adjust the pH using caustic soda.
After adjusting to 7.0 and putting it into a 30 volume jar fermenter, it was sterilized at 120Kg for 10 minutes to make the liquid volume 10. Alcaligenes No. 51 was added to the same medium composition as above prepared separately, and 0.01% yeast extract was added to the medium.
- Inoculated with Z strain (Feikoken Bacteria No. 6104) and incubated at 30°C.
20 ml of the seed culture solution cultured with shaking for 48 hours was transferred to the above main culture solution, and cultured with aeration at 30°C for 150 hours. After the culture was completed, the cells were collected using a centrifuge and the cells were classified. Separately, dissolve 186 mg of EDTA disodium dihydrate in N/10 phosphate buffer (PH7.0), and then
The volume of the solution adjusted to 7.0 was 100 ml. This solution 100
Pour the entire amount of the above bacterial cell types into ml and disperse at 30℃ and 120℃.
After holding for a while with occasional stirring,
The decomposition rate of EDTA reached 80%. Example 5 A solution containing about 1000 ppm of EDTA was prepared by diluting the waste color photographic bleach-fix solution with distilled water. Add 1 potassium phosphate to 1000ml of this solution.
6.8g, magnesium sulfate/7hydrate 0.24g, calcium chloride/dihydrate 0.5mg, manganese chloride 3mg,
After adding 10 mg of zinc sulfate heptahydrate, 500 ml of the solution whose pH was adjusted to 7.0 with sodium hydroxide was placed in a 3000 ml Sakaguchi flask to serve as the main culture solution. Separately, add Pseudomonas No. 51-Y strain (Feikoken Bacteria No. 6103) to 50 ml of a culture solution having the culture medium composition described in Example 1.
A platinum loop of the slant culture was inoculated and cultured in a reciprocating shaker at 30°C for 150 hours.The solution was aseptically collected by centrifugation and used as an inoculum. This entire amount was added to the above-mentioned main culture solution and cultured at 30°C for 150 hours using a reciprocating shaker.The decomposition rate of EDTA was measured from the remaining amount of EDTA in the supernatant and was found to be 65%.

Claims (1)

[Claims] 1. Ethylenediaminetetraacetic acid, which is characterized in that bacteria belonging to the genus Pseudomonas or bacteria belonging to the genus Alcaligenes, which have the ability to decompose ethylenediaminetetraacetic acid, are brought into contact with ethylenediaminetetraacetic acid, a metal complex of ethylenediaminetetraacetic acid, or salts of these compounds. Decomposition processing method.