JPH07107967A - New microorganism - Google Patents

Abstract

PURPOSE:To provide a microorganism which can decompose formamide, alkylformamides, alkylamines, formic acid, phenol or the like in their high concentration or in the presence of salts of high concentration. CONSTITUTION:A bacterium which belongs to Arthrobacter capable of decomposing formamide, alkylformamides, alkylamides, formic acid, phenol or the like in their high concentration or in the presence of their salts of high concentration is provided.

Description

Detailed Description of the Invention

[0001]

The present invention relates to a novel microorganism,
More specifically, amines, amides, ammonium salts, aldehydes, phenols, etc., such as dialkylformamide, monoalkylformamide, formamide, monoalkylamine, dialkylamine, trialkylamine, tetraalkylammonium salt, formic acid, formaldehyde, phenol, etc. The present invention relates to a novel bacterium that utilizes and decomposes.

[0002]

2. Description of the Related Art Among the compounds produced and used in the chemical industry and the like, dialkylformamide, monoalkylformamide, formamide, tetraalkylammonium salt, formic acid, formaldehyde are environmental pollutants. , Phenol, etc.
There are incineration methods, distillation recovery methods, and activated sludge methods as the treatment of these wastewaters. However, the incineration method and the distillation recovery method are expensive and the activated sludge method is an inefficient removal method such as a large amount of dilution because it is toxic to microorganisms.

In addition, industrial wastewater often contains inorganic salts such as sodium chloride, but these are also toxic to microorganisms at concentrations above a certain level, making treatment more difficult. There is.

For example, dialkylformamide, monoalkylformamide, formamide, monoalkylamine, dialkylamine, trialkylamine, tetraalkylammonium salt, formic acid, etc. contained in waste water should be decomposed and removed by using a degrading bacterium. The method is disclosed in JP-B-51-30151, JP-B-54-1792, JP-A-59-227294 and JP-A-64-
60371, JP-A-64-63372, JP-A-1-135595, and JP-A-1-148398.
Japanese Patent Application Laid-Open No. 2-72864, Japanese Patent Application Laid-Open No. 2-72
No. 865, JP-A-3-217298, etc., the removable concentration is 3% by weight or less for dimethylformamide and tetramethylammonium salt, and 1% by weight or less for methylamines. The resistance to inorganic salts is also low and its processing capacity is insufficient.

Further, the phenol contained in the waste water is
The method of decomposing and removing using a degrading bacterium is disclosed in JP-B-57-57199, JP-B-57-57200, JP-B-60-15397, and JP-A-3-675.
81, JP-A-4-363194, JP-A-1
No. 224094, the resistance to inorganic salts is low and the processing capacity is insufficient.

[0006]

The present inventors have found that amides, amines, ammonium salts, aldehydes or phenols contained in industrial wastewater, such as dialkylformamide, monoalkylformamide, formamide, monoalkylamine, dialkyl. Widely used in nature to search for microorganisms that have a high assimilation capacity for amines, trialkylamines, tetraalkylammonium salts, formic acid, formaldehyde, phenols, etc., and are resistant to inorganic salts such as sodium chloride. As a result, the present invention has been completed by discovering novel microorganisms in soil that effectively assimilate dimethylformamide and the like and further show resistance to inorganic salts such as sodium chloride.

Thus, the present invention relates to amides, amines, ammonium salts, aldehydes or phenols such as dialkylformamides, monoalkylformamides, formamides, monoalkylamines, dialkylamines,
Trialkylamine, tetraalkylammonium salt,
One or more of the above compounds using cells and / or treated products of novel microorganisms of the genus Arthrobacter characterized by assimilating and decomposing formic acid, formaldehyde, phenol, etc., or a mixture thereof. The present invention relates to a method for decomposing an organic compound by a microorganism, which comprises decomposing.

Of the above compounds, dialkylformamide, monoalkylformamide, monoalkylamine,
In the dialkylamine, trialkylamine, and tetraalkylammonium salt, the alkyl group is not particularly limited, but the alkyl group is preferably a compound having 1 to 6 carbon atoms. The bacterium used in the present invention belongs to the genus Arthrobacter and includes amides, amines, ammonium salts, aldehydes or phenols such as dialkylformamide, monoalkylformamide, formamide, monoalkylamine, dialkylamine, trialkylamine and tetraalkyl. There is no particular limitation as long as it is a strain capable of efficiently decomposing ammonium salts, formic acid, formaldehyde, phenol and the like.

The mycological properties of the microorganism of the present invention (hereinafter abbreviated as "the present bacterium") are as follows. The microbial accession number of this strain at the Institute of Biotechnology, Institute of Industrial Science and Technology is FER
MP-13844. The mycological properties of Arthrobacter NT-8, which is a representative strain of this bacterium, are as follows.

1. Morphology The culture was performed at 30 ° C. on a broth agar medium. (1) Shape of cells In the early stage of culture, the shape of bacilli is shown.
Then, the shape of the cocci is shown. (Has a cell cycle.) (2) Cell size In the case of rod-shaped: width 0.5 to 0.
9 μm, length 0.8-3.0 μm When cocciform: diameter 0.5-0.9 μm (3) No motility (4) No sporulation (5) Gram stain positive (6) Acid-negative

2. Growth state in each of the following media (1) Meat broth agar plate medium Degree of growth: good Properties of colony Shape: circular Surface ridge: convex circular shape. Surface shape: Smooth Edge: Smooth All edges Color: White Transparency: Opaque Gloss: Shiny

(2) Glucose broth agar plate medium Degree of growth: good Colony properties Shape: circular Surface ridge: convex circular. Surface shape: Smooth Edge: Smooth All edges Color: White Transparency: Opaque Gloss: Shiny

(3) Broth agar slant medium Growth level: good Shape: thread Surface: smooth Color: white Clarity: opaque Gloss: glossy

(4) Glucose broth agar slant medium Degree of growth: good Shape: thread Surface: smooth Color: white Clarity: opaque Gloss: glossy

(5) Liquid broth of broth with precipitate that grows on the whole liquid (6) Liquid broth of glucose broth with precipitate that grows on the whole liquid

3. Physiological properties (1) Reduction of nitrate: Positive (2) Denitrification reaction: Negative (3) MR test: Negative (4) VP test: Negative (5) Indole formation: Negative (6) Hydrogen sulfide formation: Negative (7) Starch hydrolysis: negative (8) Gelatin liquefaction: positive (9) Utilization of citric acid (in Koser's medium): Not utilized

(10) Use of inorganic nitrogen source: use both ammonium salt and nitrate (11) Dye formation: negative (12) urease: positive (13) oxidase: negative (14) catalase: positive (15) OF Fist (Hugh Raifson (Hugh
Leifson method): Negative (16) Growth range: It grows in the range of pH 5.5 to 10.5. A pH of 6-9 is desirable. It does not grow at temperatures of 5 ° C and 45 ° C. A temperature of 20 to 35 ° C is desirable. (17) Attitude toward oxygen: aerobic (18) Generation of acid and gas from sugar

Production of acid Production of gas ───────────────────────── D-glucose--D-xylose--D-fructose--Saccharose --- Maltose --- Lactose --- Starch --- Glycerin --- D-sorbitol --- ──────────────────────────

[0019]

(20) Salt tolerance: Grows in a medium containing 10% by weight NaCl. (21) Cell wall structure: contains LL-diaminopimelic acid. (22) Separation source: soil

From the above-mentioned mycological properties, BER-GEY'S MANUAL OF Sy
The second part of the Stem Bacteriology
Volume, Sneath (PETER HAASNEATH) et al., Williams and Wilkins (WILL)
According to a search by IAMS & WILKINS, (1986), this strain was identified as Arthrobacter ( Art.
bacterium ).

The isolation of this bacterium was carried out as follows. That is, DMF liquid medium (DMF per liter of distilled water
_{10g, (NH 4) 2 SO} 4 1g, KH 2 PO 4
_{0.5g, K 2 HPO 4 0.5g,} MgSO 4 · 7H
₂ O 0.2 g, NaCl 0.1 g, yeast extract 0.02 g, CaCl ₂ .2H ₂ O 2 mg, F
_{eSO 4 · 7H 2 O 2mg,} MnSO 4 · 4-5H 2 O
2 mg, ZnSO ₄ 7H ₂ O 7 mg, pH 7.0 adjusted, 121 ° C. 15 minutes wet heat sterilization) Each 5 mL of test tube with silicon stopper was inoculated with soil sampled from various places, and shake culture was performed at 30 ° C. It was

For those showing an increase in turbidity, the following four cultures were carried out under the same conditions as above. When the turbidity was increased in the fourth accumulation culture, the culture solution was changed to DMF / NaCl agar plate medium (NaCl in the DMF liquid medium was changed to 10 g / 1, and agar was added at 1.5%). Inoculated with the solidified) and 30 ℃
The present bacterium was isolated by culturing in a single colony.

For culturing the bacterium, as carbon sources, organic substances such as dialkylformamide, monoalkylformamide, formamide, mono-di-trialkylamine, tetraalkylammonium salt, formic acid, formaldehyde, ethanol, methanol, phenol, etc., Sugars such as glucose and sucrose, meat extracts, and natural products such as molasses can be used. As the nitrogen source, ammonium salts such as ammonium sulfate, nitrates such as sodium nitrate, natural products such as peptone and corn steep liquor can be used.

When a substance such as dimethylformamide which is a carbon source that can be assimilated by the bacterium and which is a nitrogen-containing compound is used as the carbon source, no other nitrogen source may be added. As the inorganic component, potassium salt,
Calcium salt, sodium salt, magnesium salt, iron salt,
Manganese salts, zinc salts, phosphates and the like can be used. Further, the trace amount of inorganic components can be substituted by using tap water. When vitamins and the like are required, corn steep liquor, yeast extract and the like can be used.

The culture temperature is 10 to 40 ° C., preferably 20.
The temperature is ˜35 ° C., the pH is 5.5 to 10.5, preferably 6.0 to 9.0, and the culture is performed aerobically. This bacterium is
Dialkylformamide, monoalkylformamide,
Formamide, monoalkylamine, dialkylamine, trialkylamine, tetraalkylammonium salt, formic acid, formaldehyde, and phenol have excellent assimilation and decomposition ability, and therefore can be used for treating wastewater containing the above organic compounds. .

The present bacterium is added to wastewater containing the above organic compound, and depending on the type of wastewater, an inorganic component such as corn steep liquor is added so that the present bacterium can grow, and the temperature is 10 to 40 ° C., preferably 20. ~ 35 ℃, pH 5.5
The organic compound is removed by keeping the temperature at 10.5, preferably 6.0 to 9.0, and performing the treatment aerobically.

The concentration of the organic compound may be within a concentration range that can be treated by the bacterium, but is usually 5% by weight for dimethylformamide, formamide, tetramethylammonium salt, trimethylamine, dimethylamine, methylamine and formic acid. Hereafter, it is preferably 3% by weight or less, and 0.1% by weight or less for formaldehyde and phenol. In addition, this bacterium is ammonium salt, potassium salt, calcium salt, sodium salt, magnesium salt,
Resistant to inorganic salts such as iron salts, manganese salts, zinc salts, and phosphates, 10 wt% NaCl or 15
Since it also grows in a medium containing wt% Na ₂ SO ₄ , the treatment is possible even when the wastewater contains a high concentration of inorganic salts.

Normally, activated sludge cannot stably treat wastewater having a high inorganic salt concentration, and the NaCl concentration is 1.0% by weight.
The above wastewater is difficult to treat, but by using this method, for example, wastewater containing 1% or more NaCl, and further 3%
It is also possible to treat the wastewater containing NaCl. In the decomposition of the organic compounds in the wastewater according to the present invention, it can be detoxified by culturing the bacterium, but it is detoxified by contacting the crushed product of the bacterial cells, the culture solution and the immobilized product of the bacterial cells with the wastewater. You can also do it. In addition, as a method for producing a crushed product of bacterial cells, ultrasonic waves may be applied to destroy the bacterial cells, or mechanically.

Specific methods of culturing or contacting under the aerobic conditions include methods such as aeration and stirring, as well as methods conventionally known in aerobic wastewater treatment, for example, a sprinkling filter method, The dipping filter method, the bacterial cell fixing method and the like can be used. As the immobilization method, a carrier binding method (physical adsorption method, ionic binding method, covalent binding method), a cross-linking method, an encapsulation method (lattice type, capsule type), or the like can be used, and the immobilization method should also be used. Representative polymer materials that can be used include polyacrylamide, κ-carrageenan, sodium alginate, agar, polyvinyl alcohol, and sodium polyacrylate.

In addition, since the waste water often contains other compounds such as dialkylformamide that can be assimilated and decomposed by the bacterium, the bacterium and the bacterium cannot be treated in such waste water. It is possible to use a combined method of bacterial cells that decomposes other compounds, or a method of further treating with activated sludge after treatment with the present bacterial cells.

[0032]

EXAMPLES The present invention will be described in more detail below with reference to examples. The present invention is not limited to the embodiments.

Example 1. Per liter of distilled water (NH
₄ ) ₂ SO ₄ 1 g, KH ₂ PO ₄ 0.5 g, K ₂ H
PO ₄ 0.5 g, MgSO ₄ .7H ₂ O 0.2 g,
NaCl 0.1 g, yeast extract 0.0
2 g, CaCl ₂ · 2H ₂ O 2 mg, FeSO ₄ · 7H
₂ O 2 mg, MnSO ₄ .4-5H ₂ O 2 mg and Z
A predetermined amount of dimethylformamide was added to a basal medium having a composition of 7 mg of nSO ₄ .7H ₂ O to adjust the pH to 7.0, and then Arthrobacter NT-8 was added to the medium subjected to heat sterilization at 121 ° C. for 15 minutes. Inoculate and incubate at 30 ℃ for 7 days,
Its assimilability was tested. The results are shown in Table 1.

[0034]

[Table 1]

Example 2. 30 g of dimethylformamide per 1 liter of the basal medium of Example 1, a predetermined amount of Na ₂ S
After adding O ₄ and adjusting the pH to 7.0, 121 ℃
Arthrobacter NT was added to the medium that had been wet sterilized for 15 minutes.
-8 was inoculated and cultured at 30 ° C for 7 days, and its assimilation ability was examined. The results are shown in Table 2.

[0036]

[Table 2]

Example 3. Na _{2 in} the medium of Example ₂
The assimilation of this bacterium was examined in exactly the same manner as in Example 2 except that NaCl was used instead of SO ₄ . The results are shown in Table 3.

[0038]

[Table 3]

Example 4. In the medium of Example 1, the assimilability of this bacterium was examined in exactly the same manner as in Example 1 except that a predetermined carbon source was used instead of dimethylformamide.
The results are shown in Table 4.

[0040]

[Table 4]

Example 5. In the medium of Example 4, the assimilation ability of the bacterium was examined in exactly the same manner as in Example 4, except that a predetermined amount of NaCl was added per liter of the basal medium.
The results are shown in Table 5.

[0042]

[Table 5]

Example 6. Suspended 1 g (wet weight) of Arthrobacter NT-8 cultured in 4 ml of 0.9% saline solution,
0.75 g of acrylamide, 40 mg of N, N'-methylenebisacrylamide, 0.5 ml of 5% β-dimethylaminopropionitrile, and 2.5% of potassium persulfate 2.5%.
5 ml was added and the mixture was allowed to stand at 20 ° C. for 30 minutes. The obtained gel was washed with 0.9% saline solution and passed through a 10 mesh sieve to obtain a fine gel. The immobilized cell gel thus obtained was added to 200 ml of the medium of Example 1 and treated at 30 ° C. for 7 days to test its assimilability. The results are shown in Table 6.

[0044]

[Table 6]

[0045]

INDUSTRIAL APPLICABILITY The microorganism of the present invention makes it possible to efficiently decompose a dialkylformamide or the like, which is difficult to treat in industrial wastewater, to a high concentration by the microorganism. It is a great invention.

Claims (5)

[Claims] 1. A novel microorganism of the genus Arthrobacter characterized by degrading at least one substance selected from the group consisting of amides, amines, ammonium salts, aldehydes and phenols. 2. The novel microorganism of the genus Arthrobacter according to claim 1, which decomposes at least one of dialkylformamide, monoalkylformamide and formamide. 3. The method according to claim 1, wherein at least one of monoalkylamine, dialkylamine, trialkylamine and tetraalkylammonium salt is decomposed.
Arthrobacter described in
r ) A novel microorganism of the genus. 4. The novel microorganism of the genus Arthrobacter according to claim 1, which decomposes at least one of formic acid, formaldehyde and phenol. 5. The novel microorganism according to any one of claims 1 to 4, wherein the microorganism belonging to the genus Arthrobacter is Arthrobacter NT-8.