Tetrahydrofuran-degrading bacteria and application thereof

Abstract

The present application relates to the technical field of microorganism and sewage treatment, and provides a tetrahydrofuran degrading bacteria and application thereof, the bacteria is Agromyces sp. HZ-O-036, and the preservation number is CGMCC No. 33509. The Agromyces sp. HZ-O-036 of the present application can degrade wastewater containing 100000 mg / L of tetrahydrofuran at most, can quickly degrade wastewater containing 2000-10000 mg / L of tetrahydrofuran within 48 h, and can adapt to the environment with pH 6-9, 30-35 DEG C, and salinity 0%-3%, and has the advantages of strong adaptability and high degradation efficiency.

Description

TECHNICAL FIELD

[0001] The present application relates to the field of microbial technology, and particularly relates to a tetrahydrofuran degrading bacterium and application thereof. BACKGROUND

[0002] Tetrahydrofuran (THF) is often used as a "universal solvent" in surface coatings, anticorrosive coatings and film coatings, inks, extractants and synthetic spandex, polyester and other materials. With the rapid development of the pharmaceutical and chemical industries, the demand and discharge of tetrahydrofuran have rapidly increased, and the resulting environmental pollution problem cannot be ignored. Studies have shown that low concentrations of tetrahydrofuran can severely affect the efficiency of activated sludge in wastewater treatment systems, resulting in a significant reduction in the treatment efficiency of the wastewater system.

[0003] The commonly used method for removing tetrahydrofuran is chemical degradation, but this method is high in cost and produces secondary pollution. Biological degradation of tetrahydrofuran is one of the most safe and reliable methods at present, and has the advantages of low cost, no secondary pollutants, and high degradation efficiency. Currently, tetrahydrofuran degrading bacteria mainly include Rhodococcus, Mycobacterium, Pseudonocardia and the like, but these bacteria generally have weak tetrahydrofuran degradation capacity and poor efficiency in wastewater treatment. Therefore, how to obtain a strain with high tetrahydrofuran degradation capacity is a problem to be solved. SUMMARY

[0004] Therefore, the present application provides a tetrahydrofuran degrading bacterium with high tetrahydrofuran degradation capacity and application thereof.

[0005] The technical scheme of the present application is implemented as follows: on the one hand, the present application provides a tetrahydrofuran degrading bacterium, which is Agromyces sp. HZ-O-036 with a preservation number of CGMCC No. 33509.

[0006] On the other hand, the present application also provides application of the tetrahydrofuran degrading bacterium in treating tetrahydrofuran wastewater.

[0007] On the basis of the above technical scheme, preferably, the Agromyces sp. HZ-O-036 is inoculated into wastewater containing tetrahydrofuran to degrade the tetrahydrofuran.

[0008] On the basis of the above technical scheme, preferably, the inoculation amount of the Agromyces sp. HZ-O-036 is 1v% to 2v%.

[0009] On the basis of the above technical scheme, preferably, the pH value of the wastewater is 6 to 9.

[0010] On the basis of the above technical scheme, preferably, the salinity of the wastewater is 0wt% to 3wt%.

[0011] Preferably, the temperature of the wastewater is 30-35 DEG C.

[0012] Preferably, the concentration of tetrahydrofuran in the wastewater is 200-10000 mg / L.

[0013] The tetrahydrofuran-degrading bacterium and the application thereof have the following beneficial effects relative to the prior art:

[0014] The soil fungus HZ-O-036 can degrade wastewater containing 100000 mg / L of tetrahydrofuran, and can quickly degrade wastewater containing 2000-10000 mg / L of tetrahydrofuran within 48 hours, and can adapt to an environment with pH 6-9, 30-35 DEG C, and salinity 0%-3%, and has the advantages of strong adaptability and high degradation efficiency. BRIEF DESCRIPTION OF DRAWINGS

[0015] In order to more clearly illustrate the technical solutions in the embodiments of the present application or the prior art, the drawings needed to be used in the embodiments or the prior art description will be briefly introduced below. Obviously, the drawings in the following description are only some embodiments of the present application, and other drawings can be obtained by those skilled in the art without creative effort on the basis of these drawings.

[0016] Figure 1 It is a scanning electron microscope photo of the soil fungus HZ-O-036;

[0017] Figure 2 It is a classification diagram of the soil fungus HZ-O-036 strain;

[0018] Figure 3 It is a diagram showing the influence of different pH on the degradation efficiency of tetrahydrofuran by the soil fungus HZ-O-036;

[0019] Figure 4 It is a diagram showing the influence of different temperatures on the degradation efficiency of tetrahydrofuran by the soil fungus HZ-O-036;

[0020] Figure 5 It is a diagram showing the influence of different salinities on the degradation efficiency of tetrahydrofuran by the soil fungus HZ-O-036;

[0021] Figure 6 It is a tetrahydrofuran degradation curve of the soil fungus HZ-O-036;

[0022] Figure 7 It is an application effect diagram of the soil fungus HZ-O-036 in high-concentration tetrahydrofuran chemical wastewater. DETAILED DESCRIPTION

[0023] The technical solutions in the embodiments of the present application will be clearly and completely described below in combination with the embodiments of the present application. Obviously, the described embodiments are only part of the embodiments of the present application, rather than all the embodiments of the present application. Based on the embodiments of the present application, all other embodiments obtained by those skilled in the art without creative work fall within the scope of protection of the present application.

[0024] Strain enrichment, separation, purification and identification of Example 1

[0025] 1. Strain enrichment

[0026] Biochemical pool activated sludge from a chemical production plant in Shanghai was used as the screening and enrichment sediment. After centrifugation of the sediment, the supernatant was removed, and the sediment was added to the enrichment medium at 10% (w / v). The enrichment was carried out at 30°C with shaking at 150 r / min. Since the carbon source in the enrichment medium only contained tetrahydrofuran, the change in COD could represent the change in the content of tetrahydrofuran. The content of COD in the medium was detected every day. Once the degradation rate of tetrahydrofuran in the medium reached more than 95%, the enrichment liquid was inoculated into new medium at a ratio of 10%, and the content of tetrahydrofuran was increased. After 4-6 weeks of acclimation according to this method, the content of tetrahydrofuran was increased from 1 g / L to 10 g / L. The tetrahydrofuran degradation efficiency of the activated sludge was further improved. At this time, the tetrahydrofuran degrading bacteria enrichment liquid was obtained, and then the enrichment bacteria liquid was separated and purified by plate dilution coating and streaking method.

[0027] Enrichment medium formula: tetrahydrofuran 1 g-10 g, Na2HPO4 1.78 g, KH2PO4 1 g, MgSO4·7H2O 0.2 g, CaCl2 0.08 g, (NH4)2SO4 0.18 g, FeSO4·7H2O 1 mg, supplemented with distilled water to 1 L, pH 7.0-7.2.

[0028] 2. Strain separation and purification

[0029] 1 ml of mixed tetrahydrofuran enrichment liquid was gradient diluted, and the sterilized purification medium was used as the diluent to dilute the enrichment liquid. The enrichment liquid was diluted to 10 -1 -10 -8 8 gradients, 100 μL of each gradient was coated on the solid purification medium, 5 parallel samples were prepared for each gradient, and the culture was incubated at 30°C for 48 h. Different colonies growing on the plates were picked and purified by streaking. After 6-7 rounds of streaking and purification, one strain was finally purified and named HZ-O-036.

[0030] Purification medium (liquid): Tetrahydrofuran 5 g, Na2HPO4 1.78 g, KH2PO4 1 g, MgSO4-7H2O 0.2 g, CaCl2 0.08 g, (NH4)2SO4 0.18 g, FeSO4-7H2O 1 mg, distilled water to 1 L, pH 7.0-7.2.

[0031] Purification medium (solid): 1.5%-2% (w / v) agar powder was added to the liquid purification medium.

[0032] 3. Strain identification:

[0033] Strain morphology: The strain HZ-O-036 was streaked on beef extract protein peptone solid medium and cultured at 30°C under aerobic conditions for 24-48 h. The colony diameter of HZ-O-036 was 0.1-0.2 mm, the colony was white, raised, smooth and non-glossy, round, smooth edge, transparent, odorless and tasteless. The micro-morphology of the strain was observed by scanning electron microscopy, and the strain was bacillus, 0.8-1.0 μm long and 0.1-0.2 μm wide, as shown in Figure 1 .

[0034] Molecular biology identification: 16S rRNA identification was performed on the strain.

[0035]

[0036] The sequencing results were compared with the known sequences in the Genbank database, as shown in the figure, strain HZ-O-036 had the highest sequence homology with Agromyces sp., and was identified as Agromyces sp.

[0037] Agromyces sp. HZ-O-036 was preserved in the China General Microbiological Culture Collection Center (CGMCC) on February 8, 2025, located at No. 1, Yihuangyuan 3rd, Beichen West Road, Chaoyang District, Beijing, China, with a postal code of 100101, and the preservation number was CGMCC No. 33509. The strain was identified as alive on February 8, 2025,

[0038] Example 2: Optimization of strain HZ-O-036 culture conditions

[0039] 1. Different initial pH

[0040] Strain activation: Take 3 mL of purified culture medium into a glass tube for sterilization. Pick a single colony on the solid enrichment medium into the liquid culture medium, and incubate at 30°C, 150 r / min on a shaker for 48 h, which is the completion of activation.

[0041] Experimental method: The activated strain was inoculated into liquid purified culture medium with different initial pH, and the inoculation ratio was 1% (v / v). The initial pH was 4.0, 5.0, 6.0, 7.0, 8.0, and 9.0, respectively, and the treatment was carried out at 30°C, 150 r / min. The COD change was detected regularly.

[0042] Experimental results: The change of COD was detected every 12 h. Since the carbon source in the culture medium was only and only tetrahydrofuran, the change of COD could represent the change of tetrahydrofuran.

[0043] The results are shown in Figure 3 and Table 1, the degradation rate can reach more than 95% between pH 6.0-9.0, and the degradation rate can reach more than 98% when the pH is 6.0-8.0, so the optimal pH for strain HZ-O-036 culture is 6.0-8.0.

[0044] Table 1: Effect of pH on degradation rate

[0045] pH 4 5 6 7 8 9 Degradation rate / % (0-48h) 25.56 85.78 99.18 99.19 98.79 96.94

[0046] 2. Effect of different temperatures on the strain

[0047] Experimental method: the activated strain was inoculated into liquid purification medium, inoculation ratio 1% (v / v), initial pH 7.0, culture temperature was set at 25℃, 30℃, 35℃, 40℃, 150r / min, and the COD was detected regularly.

[0048] Experimental results: from Figure 4 As can be seen from Table 2 and Table 2, the strain HZ-O-036 has poor tolerance to high temperature, when the temperature is 40℃, the degradation rate of tetrahydrofuran is only 20.05%. When the temperature is 30-35℃, the degradation rate of tetrahydrofuran can reach more than 92%. When the temperature decreases, the degradation rate also decreases. Therefore, the optimal culture temperature of the strain HZ-O-036 is 30-35℃.

[0049] Table 2 Influence of temperature on degradation rate

[0050] Temperature / °C 25 30 35 40 Degradation rate / % (0-48h) 51.86 99.01 92.34 20.05

[0051] 3. Influence of different salinity on the strain

[0052] Experimental method: the activated strain was inoculated into liquid purification medium, inoculation ratio 1% (v / v), initial pH 7.0, NaCl was added to the medium to make the salinity of the medium 0wt%, 1wt%, 2wt%, 3wt% and 4wt% respectively, and the culture was carried out at 150r / min, 30℃, and the COD was detected regularly.

[0053] Experimental results: from Figure 5 As can be seen from Table 3 and Table 3, when the salinity is 4wt%, the degradation rate of tetrahydrofuran by the strain HZ-O-036 is only 3.98%, almost no degradation ability. When the salinity is 0wt-3wt%, the degradation rate of tetrahydrofuran by the strain is more than 94%, and when the salinity in wastewater exceeds 1wt%, it can be considered as high salinity wastewater, therefore the strain has a wide tolerance to salinity, and can tolerate 3wt% of salt. The optimal salt concentration is 0wt%-3wt%.

[0054] Table 3 Influence of salinity on degradation rate

[0055] Salinity 0 wt% 1 wt% 2 wt% 3 wt% 4 wt% Degradation rate / % (0-48h) 99.34 99.2 98.08 94.63 3.98

[0056] 4. Tetrahydrofuran degradation curve of HZ-O-036

[0057] On the basis of the optimal strain culture pH (pH 7.0), temperature (30℃) and salinity (1wt%) selected above, the activated strain was inoculated into tetrahydrofuran liquid activation medium at a ratio of 1% (v / v), and cultured at 30℃, 150r / min, and sampled every 12h for detection.

[0058] Results are shown in Figure 6 HZ-O-036 entered the logarithmic phase after 12h of inoculation, the number of bacterial bodies increased rapidly, and the degradation rate of tetrahydrofuran also increased significantly. After 36h of culture, the strain entered the stationary phase, the concentration of bacterial bodies (OD 600 ) maintained at about 1.3, and the degradation rate of tetrahydrofuran reached 99.35%.

[0059] Example 3 Effect of strain on high-concentration tetrahydrofuran wastewater

[0060] Tetrahydrofuran wastewater: effluent from the aerobic tank of a wastewater treatment system of a chemical enterprise, the concentration of tetrahydrofuran was adjusted according to the experimental requirements.

[0061] Experimental method: the activated strain was inoculated into tetrahydrofuran wastewater with different concentrations, and the content of tetrahydrofuran in the wastewater was adjusted to 2000mg / L, 5000mg / L, 10000mg / L, 20000mg / L, 50000mg / L, and 100000mg / L, respectively. The initial pH of the wastewater was 7.2, the salinity was 1.5%, and the treatment was carried out at 30°C and 150r / min. The content of tetrahydrofuran in the wastewater was detected periodically, and the experimental results are shown in Table 4.

[0062] Table 4 Experimental results of strain HZ-O-036 treating tetrahydrofuran wastewater with different concentrations

[0063]

[0064] As shown in Table 4, tetrahydrofuran wastewater with a concentration of 100000mg / L had an inhibitory effect on the strain, and the degradation rate was 54.36%. When the concentration of tetrahydrofuran was 2000-50000mg / L, the degradation rate could reach more than 90% within 72h. When the concentration of tetrahydrofuran was 2000-10000mg / L, the degradation rate could reach more than 90% within 48h. With the increase of the concentration of tetrahydrofuran, the degradation time needs to be prolonged to reach a certain degradation rate, therefore, in practical application, the strain HZ-O-036 can quickly degrade tetrahydrofuran with a concentration of 2000-10000mg / L.

[0065] Example 4 Application examples of the strain

[0066] 1. Application of the strain in high-concentration tetrahydrofuran chemical wastewater

[0067] The high concentration tetrahydrofuran wastewater of a chemical enterprise in Shanghai is used, the COD concentration is 50-100 thousand mg / L, and the wastewater needs to be treated to below 3000 mg / L before being discharged into the sewage treatment system. A single reaction tank is set up in the plant to treat the high concentration tetrahydrofuran wastewater before being discharged into the sewage treatment system. The total volume of the stored wastewater is 3000 L, and then 300 L of high concentration tetrahydrofuran wastewater is produced every day. The pH of the wastewater is 7.0-7.2, and the salinity is 0.5%-1.5%.

[0068] The strain HZ-O-036 is added. The strain HZ-O-036 is added into the reaction tank at a ratio of 1% (v / v) every day, continuously for 5 days, and then continuously reacted for 5 days. The concentration of tetrahydrofuran is reduced from the initial 61700 mg / L to 947 mg / L, and the degradation rate is 98.46% (see Figure 7 ). The treated tetrahydrofuran wastewater can be discharged into the sewage treatment system of the enterprise. Then 300 L of high concentration tetrahydrofuran wastewater is treated every day, and the stable operation is maintained for 1 month. The COD concentration of the treated wastewater is below 3000 mg / L.

[0069] 2. Application of the strain in high-salinity tetrahydrofuran wastewater

[0070] The tetrahydrofuran wastewater produced in the production process of a chemical enterprise in Taizhou is used. The tetrahydrofuran content in the wastewater is 3000-5000 mg / L, the salinity is 1.5%-2%, and the pH is 6-8. The COD of the effluent of the plant exceeds the standard (the COD of the effluent of the plant is required to be below 500 mg / L), and the COD of the effluent is 800-1500 mg / L.

[0071] The tetrahydrofuran-degrading strain HZ-O-036 is added at a ratio of 1‰ (v / v) every day before the aerobic tank of the sewage treatment plant of the enterprise, continuously for 1 week. The change of tetrahydrofuran is detected every day. The tetrahydrofuran content in the effluent is reduced to 421 mg / L after 72 h. The effluent is continuously operated for 1 month. The effluent indexes are shown in Table 5. The COD of the effluent is maintained at 200-400 mg / L, which meets the discharge requirements.

[0072] Table 5 Application of the strain in the treatment of high-salinity tetrahydrofuran wastewater

[0073] Item pH Tetrahydrofuran content (mg / L) COD (mg / L) Influent 6-8 3000-5000 5000-6000 Effluent 7-8 40-90 200-400

[0074] The above only describes the preferred embodiments of the present application and is not used to limit the present application. Any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the present application shall be included in the protection scope of the present application.

Claims

1. A tetrahydrofuran-degrading bacterium, characterized by: The bacterial species is *Pseudomonas aeruginosa* ( Agromyces sp. HZ-O-036, with accession number CGMCC No.33509.

2. The use of the THF-degrading bacteria according to claim 1 in the treatment of THF waste water, characterized in that: The soil mold ( Agromyces sp. HZ-O-036 was inoculated into wastewater containing tetrahydrofuran to degrade tetrahydrofuran.

3. Use according to claim 2, wherein: The soil mold ( Agromyces sp. The inoculation dose of HZ-O-036 is 1v%~2v.

4. The use according to claim 2, characterized in that: The pH value of the wastewater is 6-9.

5. The use according to claim 2, characterized in that: The salinity of the wastewater is 0wt%-3wt%.

6. The use according to claim 2, characterized in that: The temperature of the wastewater is 30℃-35℃.

7. The use according to claim 2, wherein: The concentration of tetrahydrofuran in the wastewater is 200-10000mg / L.

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