Method for improving dibutyl phthalate degradation efficiency of microorganisms by utilizing biochar

Abstract

The invention relates to the technical field of new pollutant degradation, and particularly discloses a method for improving the dibutyl phthalate degradation efficiency of microorganisms by using biochar. The method comprises the following steps: fermenting straw powder by using a trichoderma asperellum T-1 spore suspension, performing pyrolysis carbonization on fermented straw residues to obtain biochar, and adding the biochar into a pollutant solution containing degrading bacteria for reaction. When the method is applied to wastewater, dibutyl phthalate pollution in water can be efficiently removed, the negative influence of dibutyl phthalate on the environment is reduced, and the water quality is improved. According to the method, the microbial degradation efficiency can be improved by 44.8%, and the method has a good application prospect.

Description

Technical Field

[0001] The present invention relates to the field of new pollutant degradation, and in particular to a method and application of utilizing biochar to improve the efficiency of microbial degradation of dibutyl phthalate. Background Art

[0002] Di-n-butyl phthalate (DBP) is a common plasticizer that has teratogenic, carcinogenic, and mutagenic hazards. It has been listed as an environmental priority pollutant and is widely used in cosmetics, plastics, paints, daily necessities and other industries to enhance the plasticity, ductility and cleanliness of products. At present, DBP has been detected in various environmental media, animals, plants, and humans around the world, becoming a type of ubiquitous and highly concentrated organic pollutant. Since DBP is not covalently bound to plastic products, but rather through physical and chemical reactions, the plastic additive DBP in plastic products can easily enter the environment.

[0003] The daily removal methods for DBP mainly include adsorption, microbial degradation and catalytic degradation. Each of these methods has its own advantages and disadvantages. The adsorption method is a physical process that can only transfer DBP but cannot remove it fundamentally. In addition, the adsorption method is not effective for organic pollutants dissolved in water. Microbial degradation uses the metabolism of microorganisms to degrade organic pollutants in wastewater, which can achieve complete removal of DBP. However, the treatment cycle is long and the requirements for water quality and environmental conditions are high. Therefore, it is particularly important to find a method that can make functional bacteria remove DBP more stably and efficiently in the environment.

[0004] Biochar has the characteristics of large specific surface area, high porosity, rich surface functional groups, good chemical stability and high mechanical stability. It is usually used for the adsorption of pollutants. The organic combination of biochar and functional strains can not only adsorb the functional bacteria and make them more stable in the environment, but also improve the degradation efficiency of the functional bacteria, so that the functional strains can remove DBP more stably and efficiently in the environment. Summary of the invention

[0005] The invention aims at the application problem of the degradation efficiency of dibutyl phthalate-degrading bacteria in sewage treatment and provides a method and application of improving the degradation efficiency of dibutyl phthalate-degrading bacteria by using biochar.

[0006] In order to achieve the above object, the present invention adopts the following technical means:

[0007] A method for improving the degradation efficiency of dibutyl phthalate-degrading bacteria by using biochar is provided, comprising the following steps:

[0008] 1) adding biochar to a degradation system solution containing dibutyl phthalate;

[0009] 2) Add dibutyl phthalate-degrading bacteria to the degradation system solution and place it in a shaker for reaction.

[0010] The concentration of dibutyl phthalate in the MSM degradation system solution containing dibutyl phthalate is 100 mg / L to 300 mg / L. Furthermore, the concentration of the MSM solution containing dibutyl phthalate is 200 mg / L.

[0011] The biochar is a kind of natural biochar or fermented biochar.

[0012] The natural biochar preparation method specifically includes: placing straw powder into a tube furnace at 4-6°C·min -1 The temperature was raised to 400-600℃ at a rate of 1-3 hours, during which nitrogen was introduced to achieve an oxygen-free environment at a rate of 4-6 L·min -1 After the temperature of the tube furnace is cooled down, the natural biochar is taken out and ground and sieved to obtain natural biochar. Further, the preparation method of natural biochar is to place the natural straw powder in a crucible and flatten it, put it into the tube furnace, and heat it at 5℃·min -1 The temperature was raised to 500 °C at a rate of 1.50 °C and pyrolyzed for 2 h. During this period, high-purity nitrogen was introduced to achieve an oxygen-free environment at a ventilation rate of 5 L min -1 When the temperature of the tube furnace drops to room temperature, take out the crucible and grind and sieve the straw charcoal.

[0013] The fermentation biochar preparation specifically includes: adding Mandel culture solution to straw powder, adding Trichoderma spinulosa T-1 spore suspension after steam sterilization, placing in an incubator at 25-30°C for 5-7 days, adding sodium citrate buffer after fermentation, placing in a shaker, shaking at 25-30°C and 100-200 rpm for 0.5-1.5 hours, passing the solid-liquid mixture through gauze, drying at 60-70°C for 40-55 hours to obtain powder, placing the powder in a tube furnace, and evaporating at 4-6°C·min -1 The temperature was raised to 400-600℃ at a rate of 1-3 hours, during which nitrogen was introduced to achieve an oxygen-free environment at a rate of 4-6 L·min -1 After the temperature of the tubular furnace cools down, take it out, grind it and sieve it to obtain fermented biochar. Furthermore, the fermented biochar preparation conditions are as follows: 30g of straw powder is placed in a conical flask, Mandel culture solution is added, and after high-pressure steam sterilization, Trichoderma spinulosa T-1 spore suspension is added, and the conical flask is placed in a 28°C incubator for 6 days. After the fermentation is completed, sodium citrate buffer is added to the conical flask, and it is placed in a shaker and shaken at 30°C and 150rpm for 1h; the solid-liquid mixture is passed through eight layers of gauze and dried at 65°C for 48h to obtain powder. The powder is placed in a tubular furnace and shaken at 5°C·min -1The temperature was raised to 500 °C at a rate of 1.5 °C and pyrolyzed for 2 h. During this period, nitrogen was introduced to achieve an oxygen-free environment at a rate of 5 L min -1 After the temperature of the tube furnace is cooled down, the biochar is taken out, ground and sieved to obtain fermented biochar. Furthermore, the solid-liquid ratio of the straw powder to the Mandel culture solution is 1 g:3.5 ml.

[0014] Further, the spore suspension of Trichoderma spinulosa T-1 was 10 6 mL -1 The concentration was added.

[0015] Furthermore, the amount of sodium citrate buffer added is 100 mL.

[0016] Furthermore, the ratio of the biochar to the MSM solution is 3 g:1 L; and the amount of biochar added is 3%.

[0017] Furthermore, the dibutyl phthalate degradation system is 20 mL.

[0018] The MSM solution is made of the following raw materials by weight based on 1L: Na 2 HPO 4 5-6 g; KH 2 PO 4 0.5~1.5g; (NH 4 ) 2 SO 4 0.02~0.06g;MgCl 2 6H 2 O 0.05~0.15g; Ca(NO 3 ) 2 ·4H 2 O 0.025~0.075g; trace element solution 0.5~1.5mL; water balance; adjust pH value to 7~8 with NaOH and HCl, sterilize at 115~130℃ high temperature steam for 10~30min;

[0019] Furthermore, the MSM culture medium is made of the following raw materials by weight based on 1L: Na 2 HPO 4 , 5.6131g; KH 2 PO 4 ,1.00g;(NH 4 ) 2 SO 4 ,0.50g;MgCl 2 6H 2 O, 0.1 g; Ca(NO 3 ) 2 ·4H 2O, 0.05g; trace elements, 1mL. Use NaOH and HCl to adjust the pH value of the inorganic salt culture medium to 7.2, divide it into conical bottles, and sterilize it at 121℃ high temperature steam for 20min.

[0020] The trace element solution is composed of the following components based on 1L: Na 2 MoO 4 ·2H 2 O0.02~0.04g; FeSO 4 7H 2 O 1~3g; MnCl 2 ·4H 2 O 0.02~0.04g; ZnSO 4 7H 2 O0.05~0.15g; H 3 BO 4 0.2~0.4g; CoCl 2 6H 2 O 0.2~0.3g; NiCl 2 6H 2 O0.01~0.03g; water remainder.

[0021] Furthermore, the trace element solution is calculated as Na in 1L. 2 MoO 4 ·2H 2 O, 0.03g; FeSO 4 7H 2 O, 2.1385 g; MnCl 2 ·4H 2 O, 0.03g; ZnSO 4 7H 2 O,0.1g;H 3 BO 4 ,0.3g;CoCl 2 6H 2 O, 0.2826 g; NiCl 2 6H 2 O; 0.02g, pH 3.5.

[0022] Trace element solution, adjust the pH value of the inorganic salt culture medium to 7.2 with NaOH and HCl, dispense into conical bottles, and sterilize at 121℃ high temperature steam for 20min.

[0023] The dibutyl phthalate-degrading bacteria are added in the form of a solution, and the OD of the dibutyl phthalate-degrading bacteria solution is 600 Further, the OD of the dibutyl phthalate degrading bacteria solution is 600 is 1.

[0024] The volume ratio of the dibutyl phthalate degrading bacteria solution to the MSM solution is 1:20 to 1:200. Further, the volume ratio of the dibutyl phthalate degrading bacteria solution to the MSM solution is 1:50.

[0025] The reaction conditions in the shaker are 20-40°C, 100-300 rpm. Further, the degradation conditions in the shaker are 30°C, 200 rpm.

[0026] The reaction time in the shaking table is 1 to 3 days. Further, the reaction time is 2 days.

[0027] Compared with the prior art, the present invention has the following advantages:

[0028] The present invention adds biochar to a dibutyl phthalate solution containing dibutyl phthalate degrading bacteria, thereby combining an adsorption method with a biodegradation method. The biochar adsorbs the degrading bacteria while adsorbing pollutants, thereby greatly improving the degradation efficiency of the degrading bacteria and having obvious advantages.

[0029] The degradation efficiency of dibutyl phthalate degradation bacteria added with biochar in the present invention can reach 99.61%, while the efficiency of phthalate degradation bacteria without adding biochar is only 68.77%. Therefore, adding biochar can improve the degradation degree of degradation bacteria; at the same time, the degradation time of pollutants of dibutyl phthalate degradation bacteria added with biochar is reduced by 60% compared with that without adding biochar. The method of the present invention is simple, efficient, energy-saving and environmentally friendly, and provides technical support for the efficient degradation and removal of DBP in sewage. BRIEF DESCRIPTION OF THE DRAWINGS

[0030] Figure 1 This is a comparison chart of the degradation efficiency of dibutyl phthalate by degradation bacteria without adding biochar and adding different concentrations of unfermented biochar;

[0031] Figure 2 This is a comparison chart of the degradation efficiency of dibutyl phthalate by degradation bacteria without adding biochar and adding different concentrations of fermented biochar; DETAILED DESCRIPTION

[0032] The following will be combined with the drawings in the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0033] The dibutyl phthalate degrading bacteria used in the present invention is of the genus Rhodococcus pyridinivorans, purchased from Guangdong Microbiological Culture Collection Center, with a culture collection number of GDMCCNO.60054.

[0034] The preparation method of natural biochar is to place natural straw powder in a crucible and flatten it, put it into a tube furnace, and heat it at 5℃·min -1 The temperature was raised to 500 °C at a rate of 1.50 °C and pyrolyzed for 2 h. During this period, high-purity nitrogen was introduced to achieve an oxygen-free environment at a ventilation rate of 5 L min -1 When the temperature of the tube furnace drops to room temperature, take out the crucible and grind and sieve the straw charcoal.

[0035] The preparation conditions of fermented biochar are as follows: 30 g of straw powder is placed in a conical flask, Mandel culture solution is added, the solid-liquid ratio of straw powder to Mandel culture solution is 1 g:3.5 ml, and after high-pressure steam sterilization, a spore suspension of Trichoderma aspergillus T-1 is added (the Trichoderma aspergillus enzyme used in the present invention uses the China General Microbiological Culture Collection Center (CGMCC), and the deposit number is CGMCC No. 9722), and the spore suspension of Trichoderma aspergillus T-1 is heated to 10 6 mL -1 The concentration was added, and the conical flask was placed in a 28°C incubator for 6 days. After the fermentation was completed, sodium citrate buffer was added to the conical flask, the amount of sodium citrate buffer added was 100mL, and it was placed in a shaker, shaken at 30°C and 150rpm for 1h; the solid-liquid mixture was passed through eight layers of gauze and dried at 65°C for 48h to obtain a powder. The powder was placed in a crucible and flattened, placed in a tube furnace, and heated at 5°C·min -1 The temperature was raised to 500 °C at a rate of 1.50 °C and pyrolyzed for 2 h. During this period, high-purity nitrogen was introduced to achieve an oxygen-free environment at a ventilation rate of 5 L min -1 When the temperature of the tubular furnace drops to room temperature, the crucible is taken out, the straw charcoal is ground and sieved to obtain fermented biochar.

[0036] MSM medium is made up of the following raw materials by weight per liter: Na 2 HPO 4 , 5.6131g; KH 2 PO 4 ,1.00g;(NH 4 ) 2 SO 4 ,0.50g;MgCl 2 6H 2 O, 0.1 g; Ca(NO 3 ) 2 ·4H 2O, 0.05g; trace element solution, 1mL. Adjust the pH value of the inorganic salt culture medium to 7.2 with NaOH and HCl, dispense into conical bottles, and sterilize at 121℃ high temperature steam for 20min.

[0037] Furthermore, the trace element solution is made of the following raw materials by weight per liter: Na 2 MoO 4 ·2H 2 O, 0.03g; FeSO 4 7H 2 O, 2.1385 g; MnCl 2 ·4H 2 O, 0.03g; ZnSO 4 7H 2 O,0.1g;H 3 BO 4 ,0.3g;CoCl 2 6H 2 O, 0.2826 g; NiCl 2 6H 2 O; 0.02g, pH 3.5.

[0038] Trace element solution, 1 mL, adjust the pH value of the inorganic salt medium to 7.2 with NaOH and HCl, dispense into conical bottles, and sterilize at 121℃ high temperature steam for 20 min.

[0039] Furthermore, the volume ratio of the dibutyl phthalate degrading bacteria solution to the MSM solution is 1:50.

[0040] The experimental methods in the following examples, unless otherwise specified, are conventional methods and are performed according to the techniques or conditions described in the literature in the art or according to the product instructions.

[0041] Unless otherwise specified, the materials and reagents used in the following examples can be obtained from commercial sources.

[0042] Example 1

[0043] This embodiment provides a method for improving the efficiency of microbial degradation of dibutyl phthalate by using biochar, comprising the following steps:

[0044] The degradation experiment was carried out in batches. A clean 50 mL glass conical flask was prepared, 20 mL of MSM solution was added, and the solution was sterilized at 121 °C for 20 min before use. Methanol was used to prepare DBP mother solution for use. A final concentration of 200 mg·L -1200 μL dibutyl phthalate solution (filter sterilized), 200 μL dibutyl phthalate degrading bacteria solution (OD of dibutyl phthalate degrading bacteria solution 600 1); without adding biochar, put into shaker, 30℃, 200rpm and culture for 2 days.

[0045] Example 2

[0046] This embodiment provides a method for improving the efficiency of microbial degradation of dibutyl phthalate by using biochar, comprising the following steps:

[0047] The degradation experiment was carried out in batches. A clean 50 mL glass conical flask was prepared, 20 mL of MSM solution was added, and the solution was sterilized at 121 °C for 20 min before use. Methanol was used to prepare DBP mother solution for use. A final concentration of 200 mg·L -1 200 μL dibutyl phthalate solution (filter sterilized), 200 μL dibutyl phthalate degrading bacteria solution (OD of dibutyl phthalate degrading bacteria solution 600 1); add 0.06g (3%, m / v, that is, the ratio of natural biochar to MSM solution is 3g / L) natural biochar, put it in a shaker, 30°C, 200rpm and culture for 2d.

[0048] Example 3

[0049] This embodiment provides a method for improving the efficiency of microbial degradation of dibutyl phthalate by using biochar, comprising the following steps:

[0050] The degradation experiment was carried out in batches. A clean 50 mL glass conical flask was prepared, 20 mL of MSM solution was added, and the solution was sterilized at 121 °C for 20 min before use. Methanol was used to prepare DBP mother solution for use. A final concentration of 200 mg·L -1 200 μL dibutyl phthalate solution (filter sterilized), 200 μL dibutyl phthalate degrading bacteria solution (OD of dibutyl phthalate degrading bacteria solution 600 1); add 0.06 g (3%, m / v) fermented biochar, put into a shaker, and culture at 30°C and 200 rpm for 2 days.

[0051] Performance Testing

[0052] The group without biochar addition was used as the experimental group, and Examples 1, 2, and 3 were used as the control group. The difference between the experimental group and the control group was whether biochar was used, whether the biochar used was subjected to different pyrolysis temperatures, different amounts, and whether it was fermented.

[0053] Test results such as Figure 1As shown in the results, adding biochar to the degradation system can effectively improve the efficiency of functional bacteria in degrading DBP. The degradation efficiency of DBP-degrading bacteria in the group with biochar added was increased by 44.8% compared with that in the group without biochar added, and the degradation time was reduced by 60%. It is worth noting that the degradation efficiency of DBP-degrading bacteria in the group with fermented biochar added was increased by 9.5% compared with that in the group with unfermented biochar added.

[0054] The above descriptions are merely embodiments of the present invention and are not intended to limit the patent scope of the present invention. Any equivalent structure or equivalent process transformation made using the contents of the present invention specification, or directly or indirectly applied in other related technical fields, are also included in the patent protection scope of the present invention.

Claims

1. A method for improving the efficiency of microbial degradation of dibutyl phthalate using biochar, characterized in that: The following steps are involved: 1) adding dibutyl phthalate to be degraded into the MSM solution to form an MSM degradation system solution containing dibutyl phthalate; 2) Add biochar and dibutyl phthalate-degrading bacteria to the MSM degradation system solution and place it in a shaker for reaction.

2. The method according to claim 1, characterized in that In step 1), the concentration of dibutyl phthalate in the MSM degradation system solution containing dibutyl phthalate is 100 mg / L to 300 mg / L.

3. The method according to claim 1, characterized in that In step 2), the biochar is one of natural biochar and fermented biochar.

4. The method according to claim 3, characterized in that In step 2), the natural biochar preparation specifically includes: The straw powder was placed in a tube furnace at 4-6℃·min -1 The temperature was raised to 400-600℃ at a rate of 1-3 hours, during which nitrogen was introduced to achieve an oxygen-free environment at a rate of 4-6 L·min -1 After the temperature of the tubular furnace is cooled down, the biochar is taken out, ground and sieved to obtain natural biochar.

5. The method according to claim 3, characterized in that: In step 2), the fermentation biochar preparation specifically includes: Mandel culture medium was added to the straw powder, and after steam sterilization, a spore suspension of Trichoderma aspergillus T-1 was added, and the powder was placed in a 25-30°C incubator for 5-7 days. After the fermentation was completed, sodium citrate buffer was added, and the powder was placed in a shaker, and the mixture was shaken at 25-30°C and 100-200 rpm for 0.5-1.5 hours. The solid-liquid mixture was passed through gauze and dried at 60-70°C for 40-55 hours to obtain a powder. The powder was placed in a tube furnace and heated at 4-6°C·min -1 The temperature was raised to 400-600℃ at a rate of 1-3 hours, during which nitrogen was introduced to achieve an oxygen-free environment at a rate of 4-6 L·min -1 After the temperature of the tubular furnace is cooled down, the biochar is taken out, ground and sieved to obtain fermented biochar.

6. The method according to claim 1, characterized in that In step 2), the dibutyl phthalate-degrading bacteria are added in the form of a solution, and the OD of the dibutyl phthalate-degrading bacteria solution is 600 It is 0.8~1.

2.

7. The method according to claim 5, characterized in that In step 2), the ratio of the biochar to the MSM solution is 2-4 g: 1 L; The volume ratio of the dibutyl phthalate degrading bacteria solution to the MSM solution is 1:20 to 1:

200.

8. The method according to claim 1, characterized in that In step 1), the MSM solution is made of the following raw materials by weight based on 1L: Adjust the pH value to 7-8 with NaOH and HCl, and sterilize with high-temperature steam at 115-130°C for 10-30 minutes; The trace element solution is composed of the following components based on 1L:

9. The method according to claim 1, characterized in that: In step 2), the reaction conditions in the shaking table are 20-40° C. and 100-300 rpm.

10. The method according to claim 1, characterized in that In step 2), the reaction time in the shaking table is 1 to 3 days.

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