KR100493704B1 - Novel Trichosporon loubieri Variant and Method for Treatment of High Concentrate Waste Water Using the Same - Google Patents

Abstract

The present invention relates to a novel variant of Trichosporon loubieri resistant to high concentration wastewater, a wastewater treatment spawn agent comprising the same, and a wastewater treatment method using the spawn agent, and more specifically, to a high concentration wastewater. tri course isophorone ruby Erie SP2 having resistance (KFCC - 11319 No.), and the waste water (ⅰ) tri course isophorone ruby Erie SP2 alone, or (ⅱ) Bacillus pumil Ruth (Bacillus pumilus), Bacillus MEGATHERIUM (Bacillus megaterium) And biological pretreatment with a wastewater treatment spawn comprising at least one Bacillus bacterium selected from the group consisting of Bacillus subtilis and tricosporon rubieri SP2.

Description

Novel Trichosporon loubieri Variant and Method for Treatment of High Concentrate Waste Water Using the Same}

The present invention relates to a novel trichosporon loubieri mutant resistant to high concentration wastewater, a wastewater treatment spawn comprising the same and a wastewater treatment method for biological pretreatment in a sump using the spawn.

In general, the activated sludge process, which is a biological wastewater treatment method, is used to treat wastewater by diluting the wastewater concentration to 1500 mg / l or less based on chemical oxygen demand (COD). Wastewater treatment methods can be roughly divided into physical treatment methods, chemical treatment methods, and biological treatment methods. Of these, biological treatment is an active method of oxidatively decomposing organic matter contained in industrial wastewater, livestock wastewater, etc. using microorganisms. It is divided into aerobic treatment methods such as sludge method and biological method and anaerobic treatment methods such as oxidation paper method.

As a biological wastewater treatment method using a microbial agent, there is a method of stably treating sludge generated during the manufacture of polymer fibers using a mixed strain of Pseudomonas genus, plasil bacteria, etc. (Korean Patent Application Publication No. 1995-26824) ), A method of using mixed cultures of soil microorganisms such as genus Bacillus, Staphylococcus , Pseudomonas, Nitrosomonas , and Flavobacterium ( Flavobacterium ). 1994-21719).

In addition, spawning agents for wastewater treatment that do not require the addition of nitrogen sources and phosphoric acid, which are inevitably used in biological wastewater treatment, are known (Korean Patent Application Publication No. 2002-0039110), and various microorganisms using Pseudomonas sp. Methods for biological treatment of wastewater of chlorinated phenolic compounds of the kind are known ( Kor. J. Appl. Microbial. Biotechnol. Vol. 29, No. 2, 115-120 (2001)). Kor. J. Appl. Microbial. Biotechnol. Vol. 22, No. 2, 197-202 (1994) disclose a method for treating high concentration livestock wastewater using Spirulina platensis , which has a removal efficiency of about 50% of nitric acid and phosphate . J. Appl. Microbial. Biotechnol. Vol. 26, No. 1, 76-82 (1998) described the concentrations of COD, BOD and n-hexane extracts from wastewater treatment using Pseudomonas chlororapihis for biological treatment of oil-containing wastewater. The removal rates averaged 92.9%, 94.8% and 98.0%, respectively, and it is reported that algae ( Chlorella valgaris ) can efficiently remove nitrogen and phosphorus from wastewater.

Also, Kor. J. Appl. Microbial. Biotechnol. Vol. 27, No. 2, 153-158 (1999) showed that nitrogen and phosphorus removal rates were 43.9% and 41.7%, respectively, when Botryococcus braunii , which is known to have the highest intracellular accumulation of hydrocarbons in microalgae, was cultured. It is starting to be, Kor. J. Appl. Microbial. Biotechnol. Vol. 24, No. 6, 738-742 (1996), reported that the treatment efficiency of organic wastewater using Rhodo pseudomonas capsulata , a red, non-sulfur photosynthetic bacterium, showed more than 90% treatment efficiency. .

A method of biological purification of contaminated soil using tricosporon rubyeri is known (Korean Patent Application Publication No. 2002-444), and a method of treating wastewater or waste gas using a new tricosporon rubyeri is known (Korean patent application Publication 2002-84756). The method is characterized by passing waste water or waste gas through a carrier or filter on which a tricosphorone rubieri strain that degrades toxic and hardly decomposable chemicals is immobilized.

On the other hand, the development of resistant strains have been limited to the strains used in some medicines, and Korean Patent No. 128856 is a variant of Bifidobacterium bifidum , which has resistance to rifampicin and fluoroquinolone antibiotics. A novel variant, B. bifidum B. bifidum OFR9 (KCTC 8640P), is disclosed. The patent document discloses that the growth of Shigella dysenterial ATCC 975C is inhibited in a medium in which B. Bifidum OFR9 culture medium and BL-liquid medium are mixed to adjust the pH to 4.92. It is starting. In addition, regarding the development of resistant growth strains, Microbial. Immumol. Vol. In 21 (2) and 85-97 (1997), data was reported that resistance was induced as shown in Table 1 by inducing resistance of Enterococcus f aecalis strains.

Antibiotic Class MIC (minimum growth retardant concentration) (µg / ml) Mother strain Resistant strains Streptomycin 40 10,000 Chloramphenicol 5 500 Tetracycline 5 120 Erythromycin 10 1,000 Aminobenzyl penicillin 5 200 Penicillin G 10 200 Cephalexin 25 300 Gentamicin 5 20 Lincomycin 10 100 Kanamycin 100 10,000

In addition, rifampicin described in Korean Patent No. 280366 and Enterococcus faecails BR2, a novel mutant with various antibiotic resistance, induced 70-fold resistance of streptomycin than parent strain, erythromycin 100-fold, kanamycin Resistance was induced at 50-fold and rifampicin at 16-fold.

On the other hand, the conventional wastewater treatment agent is added to a high concentration of wastewater, that is, a high concentration of 10,000-20,000 mg / l based on chemical oxygen demand (COD) measurement, most of the seeds are killed within a few hours, and as a spawning agent of the concept of biological pretreatment, There is a problem that cannot be used. Therefore, there is a need in the art for the development of spawning agents that are resistant to high concentrations of wastewater.

The present inventors have made intensive efforts to develop a spawning agent that is resistant to wastewater containing high concentrations of organic matter. As a result, the present inventors have succeeded in preparing a new mutant of Trichosporon loubieri , and used high concentration wastewater in a sump tank . The present invention has been completed by confirming that the wastewater treatment can be carried out with improved efficiency when the wastewater treatment is carried out by biological pretreatment.

Accordingly, it is an object of the present invention to provide a trichosporon loubieri novel mutant that is resistant to high concentration wastewater.

Another object of the present invention is to provide a wastewater treatment spawning agent comprising the tricosphoron rubies novel mutant strain.

It is another object of the present invention to provide a wastewater treatment method for biological pretreatment in a sump.

Other objects and advantages of the present invention will become apparent from the following detailed description, claims and drawings.

According to one aspect of the present invention, the present invention provides a novel variant of Trichosporon loubieri SP2 (Accession No. KFCC-11319), which is resistant to high concentrations of wastewater.

The novel variant strains of the present invention are resistant to wastewater with high concentrations, ie high organic concentrations. As used herein, the term "high concentration" refers to the concentration of high organic matter, unless otherwise specified, and the high concentration is 10,000-30,000 mg / l, preferably 15,000-20,000 mg / l, most preferably based on chemical oxygen demand measurement. Preferably about 20,000 mg / l. The novel mutant strains of the present invention exhibit normal growth patterns in high concentrations of organic wastewater, such as chemical-related facility wastewater and livestock wastewater, and are very useful as a spawning agent for the treatment of high concentration wastewater.

According to another aspect of the invention there is provided (ⅰ) tri course isophorone ruby Erie (Trichosporon loubieri) SP2 alone, or (ⅱ) Bacillus pumil Ruth (Bacillus pumilus), Bacillus MEGATHERIUM (Bacillus megaterium) and Bacillus interference Tilly A wastewater treatment spawn comprising at least one Bacillus bacterium selected from the group consisting of Bacillus subtilis and Trichosporon loubieri SP2 is provided.

When treating high concentrations of wastewater according to the activated sludge method generally used for wastewater treatment, it is generally first diluted to 1500 mg / l or less based on chemical oxygen demand measurement. The spawn agent of the present invention may be introduced into a sump collecting high concentration wastewater to reduce the concentration of organic matter in the wastewater to an appropriate level, thereby greatly increasing the efficiency of the activated sludge wastewater treatment system.

The seed agent of the present invention may also be composed of Trichosporon loubieri SP2 alone, but preferably Bacillus pumilus , Bacillus megaterium and Bacillus subtilis subtilis ) is a complex seed agent comprising at least one Bacillus bacterium and Trichosporon loubieri SP2 selected from the group consisting of the most preferred Bacillus species and tricosporon rubies (3). Trichosporon loubieri ) is a complex seed containing SP2.

According to another aspect of the invention there is provided (ⅰ) tri course isophorone ruby Erie (Trichosporon loubieri) SP2 alone, or (ⅱ) Bacillus pumil Ruth (Bacillus pumilus), Bacillus MEGATHERIUM (Bacillus megaterium) to the waste water, and A wastewater treatment method comprising the step of biological pretreatment in a sump with a wastewater treatment spawn comprising at least one Bacillus bacterium selected from the group consisting of Bacillus subtilis and Trichosporon loubieri SP2 to provide.

As used herein, the term "wastewater treatment" means purifying water by removing contaminants (eg, soluble organics, suspended solids, colloidal materials, inorganics, etc.), especially organics, in the wastewater, unless otherwise specified.

In the method of the present invention, the spawning agent used may be composed of tricosphoron rubyeri SP2 alone, but preferably at least one selected from the group consisting of Bacillus pumilus, Bacillus megaterium and Bacillus subtilis. A bacterium comprising a Bacillus bacterium and Tricosporon rubieri SP2, and most preferably, a bacterium of three Bacillus species described above and a complex spawn comprising Tricosporon rubieri SP2.

The method of the present invention can be applied to various biological treatment methods, for example, it can be applied to activated sludge method, water trickling method, oxidation paper method, rotational disc method, catalytic oxidation method, etc., but most preferably applied to activated sludge method. The activated sludge method may be classified into a stage aeration method, tapered aeration method, contact stabilization method, long time aeration method, oxidation sphere method, Klaus method, pure oxygen activated sludge method and the like.

When the method of the present invention is applied to the activated sludge process, it comprises the steps of: (a) the waste water (ⅰ) tri course isophorone ruby Erie (Trichosporon loubieri) SP2 alone, or (ⅱ) Bacillus pumil Ruth (Bacillus pumilus ), a collection tank with wastewater treatment spawn comprising Trichosporon loubieri SP2 and at least one Bacillus bacterium selected from the group consisting of Bamilus megaterium and Bacillus subtilis Biological pretreatment in; (b) aeration of the pretreated wastewater in an aeration tank; (c) precipitating sludge in the aerated waste water in a settling tank; (d) returning a portion of the precipitated sludge and discarding the remaining portion; And (e) obtaining the supernatant of the final treated wastewater.

The organic matter concentration of the wastewater treated by the method of the present invention is not limited, but is preferably 10,000-30,000 mg / l, more preferably 10,000-20,000 mg / l, and most preferably 15,000 based on chemical oxygen demand measurement. -20,000 mg / l. The chemical oxygen demand herein means a value measured by various methods known in the art, and most preferably means a value measured by the potassium dichromate method.

The biological pretreatment in the sump with the spawning agent comprises 10-40%, preferably 20-35%, more preferably 25-30%, and most preferably about 30% organic matter concentration in the wastewater based on chemical oxygen demand measurement. Decrease.

Reducing the organic concentration of the high concentration wastewater by the spawning agent can greatly increase the overall efficiency of the activated sludge wastewater treatment system.

The aeration method in the aeration step may be carried out in various ways, it may be carried out by intermittent aeration or continuous aeration, etc., preferably in a continuous aeration method. According to a preferred embodiment of the present invention, the aeration step is not additionally added to the seed for decomposition of organic matter.

After this treatment, all of the above-mentioned strains survive in the activated sludge returned to the aeration tank, which is a factor for greatly improving the efficiency of the method of the present invention.

When the method of the present invention is carried out according to the above-described activated sludge process, the device for implementing the method can be largely composed of three parts: (i) a water collecting tank in which the pretreatment with the spawning agent is performed; A conveying facility for conveying part of the aeration tank and (i) active sludge.

Hereinafter, the present invention will be described in more detail with reference to Examples. These examples are only for illustrating the present invention in more detail, it will be apparent to those skilled in the art that the scope of the present invention is not limited by these examples in accordance with the gist of the present invention. .

Example 1: Strain Isolation

After separating 37 kinds of surviving strains from high concentration wastewater (more than 5,000 mg / l based on COD quantification method) in pharmaceutical wastewater treatment plant, nutritional broth 0.5%, NH ₄ NO ₃ 0.28%, KH ₂ PO ₄ 0.25%, glucose 2.0%, In a 2% glucose agar plate containing 0.0001% CaCl ₂ , 0.01 mg / l FeSO ₄ .7H ₂ O, 0.08 mg / l ZnSO ₄ , 0.02 mg / l MnSO ₄ , 0.01 mg / l CuSO ₄ and 2% agar. The plate was incubated for 2-8 days in a 15-35 ° C. incubator. Single colonies were isolated from the colonies resulting from the culture and stored.

Example 2: Selection of Effective Strains

In a Erlenmeyer flask containing 500 ml of a prepared 2% glucose liquid medium, 37 strains isolated in Example 1 were each inoculated and incubated at 15-35 ° C. for 2-8 days. This strain was inoculated with 250 ml of the spawn seed cultured per 10 liters of wastewater in a high concentration wastewater of 5,000 mg / l on the basis of COD quantification, and then treated with DO (dissolved oxygen) at 4 ° C at 4 ° C for 4 days. COD was measured. Among them, four strains decomposing more than 30% of COD were selected and isolated and preserved in 2% glucose agar plate medium. The four microorganisms selected were Trichosporon loubier , Bacillus pumilus , Bacillus megatreium and Bacillus subtilis .

Example 3 Induction of High Concentration Wastewater Resistance

Four effective strains selected in Example 2 were incubated in 2% glucose liquid medium at 15-35 ° C. for 2-8 days, and then 250 ml of the culture solution was inoculated into 10 l of 8,000 mg / l wastewater on a COD quantitative basis. It was. Subsequently, surviving strains were reacted with aeration for 4 days at 27 ° C. and passaged in 2% glucose agar plate medium. In this way resistance was induced with increasing wastewater concentrations up to 10,000 mg / l on a COD quantitative basis. The microbiological characteristics of Bacillus permilus, Bacillus megaterium and Bacillus subtilis among the microorganisms induced resistance to high concentrations of wastewater were investigated as shown in Table 2 below:

division Bacillus megarterium Bacillus pumilus Bacillus subtilis Colony Oval Oval, yellow Round, irregular cream or brown Fungus Stick Stick Stick Width (μm) 1.0 to 1.2 0.6 to 0.7 0.7 to 0.8 Bacterial Length (㎛) 3 to 5 2-3 2-3 Oxidation / fermented starch + - + Casein + + + Hypurate ± + - Gram Dyeing + + + Growth temperature (℃) 35 to 45 45-50 45 to 55 Minimum growth temperature (℃) 3 to 20 5 to 15 5-20 Catalase + + + growth Aerobic Aerobic Aerobic pH range 5 to 10 6-10 4 to 10 Salt Resistance (% NaCl) 10 7.5 10 Protease ++ ++ +++ Amylase + + + cellulose ++ +++ ++ Lipase + ++ +++

Example 4 Tricosporon Rubieri Mutant Isolation

Among the microorganisms in which resistance to high concentration wastewater was induced in Example 3, trichosporon loubieri was subjected to increased slope passage culture to induce higher wastewater resistance. First, the 2% glucose agar medium maintained at 50 ° C. was poured into a container and solidified as shown in FIG. 1 as a slope, and when the COD was measured instead of distilled water when preparing 2% glucose agar medium, it was 20,000 mg / l. After putting the agar melted, and was replaced with a medium on the slope replacement medium as shown in FIG. After solidification, Trichosporon loubieri was plated thereon , followed by incubation at 15-35 ° C. for 2-8 days to isolate and preserve the bacteria growing in the thick layer with high concentration of wastewater. The same procedure as described above was repeated 15 times to induce high concentrations of wastewater resistant bacteria. The result was a strong mutant strain that survives high concentrations of 20,000 mg / l wastewater when COD is measured. Whether the variant strain was new or not was confirmed by 28S rDNA D ₁ / D ₂ sequencing and the sequence is described in SEQ ID NO: 1.

The resulting high concentration of wastewater resistant induced spawn was named " Trichosporon loubieri SP2" and was deposited on December 28, 2002 at the Korea Microorganism Conservation Center and deposited with KFCC-. 11319 was granted.

The microbiological characteristics of Trichosporon loubieri SP2 of the present invention are shown in Table 3 below:

division Trichosporon loubieri SP2 Colony cream color Fungus Enterococci Width (μm) 4-8 μm Phenolic Compound Degradability + Glucose fermentation - Maltose - Sucrose - Huh / Jin Hypha + Lush spores + Growth temperature (℃) 40 ℃ Minimum growth temperature (℃) 10 ℃ Diphenol oxidase - Urease ± pH range 3-10 Hemi-cellulose +

Example 5 Chemical Wastewater Treatment Efficiency Measurement

In order to measure the efficiency of the chemical wastewater treatment of the spawning agent according to the present invention, four resistant toxins isolated purely in Examples 3 and 4 were respectively inoculated into a Erlenmeyer flask containing 500 ml of a 2% glucose liquid medium prepared in advance. Incubated at 15-35 ° C. for 2-8 days. After mixing the culture medium of the four resistant bacteria was inoculated 250ml per 10L of high concentration wastewater of 18,000 mg / ℓ on the basis of COD quantification, and then treated at 27 ℃ for 4 days. The results are shown in FIG. Samples were taken immediately after inoculation and the COD was measured to be 18,000 mg / L. After one day, the COD decreased by 33%, and after four days, no more than that.

As can be seen in Figure 4, after measuring the number of viable bacteria in the waste water was measured immediately after inoculation, the number of bacteria was measured, the result was reduced from 1 × 10 ⁷ / ㎖ 4 days after 1 × 10 ⁴ / ㎖. 5 is a measurement of dissolved oxygen consumption, the dissolved oxygen amount increased by 39% after one day and 21% after four days. FIG. 6 is MLSS (float concentration), increased to 250 mg / l 3 days after inoculation.

Example 6 Livestock Wastewater Treatment Efficiency Measurement

In order to measure the livestock wastewater treatment efficiency of the spawning agent according to the present invention, each of the Erlenmeyer flasks containing 500 ml of 2% glucose liquid medium prepared in advance was inoculated and incubated at 15-35 ° C. for 2-8 days. After mixing the culture solution and inoculating 250 ml per 10 l of the high concentration livestock wastewater having a COD of 13,500 mg / l, the result was measured for 9 days at room temperature. As can be seen in Figure 7, the sample was taken immediately after inoculation, and the COD was measured as a result of 13,500mg / ℓ, 42.2 days after the decrease was 22.2%. Figure 8 shows the number of viable bacteria in the wastewater, which was measured immediately after inoculation 8 x 10 ⁶ / ㎖ but increased to 3 × 10 ⁸ after 3 days, 2 x 10 ⁷ / Survival to ml.

Example 7 Measurement of Single Strain Wastewater Treatment Efficiency

In order to measure the wastewater treatment efficiency of each of the single strains of the spawning agent according to the present invention, four resistant strains isolated purely in Examples 3 and 4 were added to each of the Erlenmeyer flasks containing 500 ml of 2% glucose liquid medium prepared in advance. Inoculation was incubated at 15-35 ° C. for 2-8 days. 250 ml of this culture was inoculated into 10 l of a high concentration wastewater of 17,500 mg / l on a COD quantitative basis and then treated at 27 ° C. for 4 days, and the results are shown in FIG. 9. Samples taken immediately after inoculation and measured for COD were 16,800 mg / l of Trichosporon loubieri and increased after 1 day, 7% after 4 days, Bacillus pumilus. Bacillus pumilus was degraded to 9.8%, Bacillus megaterium 8.1%, and Bacillus subtilis 5.2%.

Having described the specific part of the present invention in detail, it is apparent to those skilled in the art that the specific technology is merely a preferred embodiment, and the scope of the present invention is not limited thereto. Thus, the substantial scope of the present invention will be defined by the appended claims and equivalents thereof.

As described in detail above, the present invention provides a trichosporon loubieri novel mutant resistant to high concentration wastewater. In addition, the present invention provides a wastewater treatment spawn agent comprising the tricosporon rubieri novel mutant strain and a wastewater treatment method using the same. Tricosporon rubies novel mutant strain of the present invention has a strong resistance to high concentration wastewater, the wastewater treatment method of the present invention using the same provides a system that can efficiently treat high concentration wastewater.

BRIEF DESCRIPTION OF THE DRAWINGS Fig. 1 shows a sloped solid medium used for the preparation of Trichosporon loubieri SP2 of the present invention;

2 is a diagram showing a sloped solid medium used in the preparation of Trichosporon loubieri SP2 of the present invention;

3 is a graph showing wastewater treatment efficiency by the method of the present invention;

4 is a graph showing a change in the number of bacteria in the wastewater during the wastewater treatment by the method of the present invention;

5 is a graph showing a change in the amount of dissolved oxygen in wastewater during wastewater treatment by the method of the present invention;

6 is a graph showing the change of suspended solids concentration in the treatment of wastewater by the method of the present invention;

7 is a graph showing the livestock wastewater treatment efficiency by the method of the present invention;

8 is a graph showing the change in the number of bacteria in the waste water during livestock wastewater treatment by the method of the present invention; And

Figure 9 is a graph showing the wastewater treatment efficiency of each strain used in the present invention.

<110> B & B Korea Co., Ltd. <120> Novel Trichosporon loubieri Variant and Method for Treatment of High Concentrate Waste Water Using the Same <130> 03DP0024 <160> 1 <170> KopatentIn 1.71 <210> 1 <211> 637 <212> DNA <213> Trichosporon loubieri SP2 <400> 1 gcatatcaat aagcggagga aaagaaacta acaaggattc ccttagtaac ggcgagtgaa 60 ccgggaagag ctcaaatttg aaatctggct gtcttcgata gtccgagttg taatctatag 120 aagcgttttc cgtgctgaac tgtgtctaag tcccttggaa cagggtatca aagagggtga 180 caatcccgta cttgacacaa tcatcagtgc tctgtgatac gctctctacg agtcgagttg 240 tttgggaatg cagctcaaaa tgggtggtaa attccatcta aagctaaata ttggcgagag 300 accgatagcg aacaagtacc gtgagggaaa gatgaaaagc actttggaaa gagagttaaa 360 cagtacgtga aattgttgaa agggaaacga ttgaagtcag tcgtgttcat tggattcagc 420 cggtcttcgg tgtacttcct ttgaacgggt caacatcagt tttgcccggt ggataaaggc 480 agtaggaatg tggcctcttc ggaggtgtta tagcctattg tcacatacac tgggggagac 540 tgaggactgc agctcgcctt ttggccgggg ttcgcccacg ttcgagctta ggatgttgac 600 ataatggctt taaacgaccc gtcttgaaac acggacc 637

Claims (7)

Trichosporon loubieri SP2 (KFCC-11319) resistant to wastewater containing high concentrations of organic matter. (Ⅰ) tri course isophorone ruby Erie (Trichosporon loubieri) SP2 alone, or (ⅱ) Bacillus pumil Ruth (Bacillus pumilus), Bacillus MEGATHERIUM (Bacillus megaterium) and B. subtilis selected from the group consisting of (Bacillus subtilis) A wastewater treatment spawn comprising one or more Bacillus bacteria and Trichosporon loubieri SP2. The waste water (ⅰ) from the group consisting of tri course isophorone ruby Erie (Trichosporon loubieri) SP2 alone, or (ⅱ) Bacillus pumil Ruth (Bacillus pumilus), Bacillus MEGATHERIUM (Bacillus megaterium) and B. subtilis (Bacillus subtilis) A wastewater pretreatment method comprising biologically pretreatment with a wastewater treatment spawn comprising at least one Bacillus bacterium and Trichosporon loubieri SP2 selected. Wastewater treatment method comprising the following steps: (a) to the waste water (ⅰ) tri course isophorone ruby Erie (Trichosporon loubieri) SP2 alone, or (ⅱ) Bacillus pumil Ruth (Bacillus pumilus), Bacillus MEGATHERIUM (Bacillus megaterium) and B. subtilis (Bacillus subtilis) Pretreating a wastewater treatment spawn comprising at least one Bacillus bacterium selected from the group consisting of and Trichosporon loubieri SP2; (b) aeration of the pretreated wastewater in an aeration tank; (c) precipitating sludge in the aerated waste water in a settling tank; (d) returning a portion of the precipitated sludge and discarding the remaining portion; And (e) obtaining the supernatant of the final treated wastewater. 4. The method of claim 3, wherein the concentration of organic matter in the wastewater is 10,000-20,000 mg / l based on chemical oxygen demand measurement. The method of claim 3, wherein the biological pretreatment with the spawning agent reduces the concentration of organic matter in the wastewater by 10-40% based on chemical oxygen demand measurement. The wastewater treatment method according to claim 4, wherein the aeration step is not additionally added with a spawning agent for decomposing organic matter.