Application of 4-methyl-1-piperazine ethylamine as nitrification inhibitor and nitrification inhibition method

Abstract

The invention provides application of 4-methyl-1-piperazine ethylamine as a nitrification inhibitor and a nitrification inhibition method, and relates to the technical field of soil treatment. Specifically, the invention relates to application of 4-methyl-1-piperazine ethylamine as a nitrification inhibitor in soil treatment and / or sewage treatment and inhibition of nitrosation activity of ammonia oxidizing bacteria. The method for inhibiting nitrification comprises the step of treating soil or water containing ammonium nitrogen and ammonia oxidizing bacteria by using 4-methyl-1-piperazine ethylamine. The concentration of the 4-methyl-1-piperazine ethylamine is not lower than 0.01 per thousand w / v by taking the volume of the soil or the water body as 100%. According to the invention, the nitrification inhibition activity of 4-methyl-1-piperazine ethylamine is found, so that the variety selection of nitrification inhibitors is enriched.

Description

Technical Field

[0001] The invention relates to the technical field of soil treatment, and in particular to application of 4-methyl-1-piperazineethylamine as a nitrification inhibitor and a method for inhibiting nitrification. Background Art

[0002] Nitrification refers to the process of ammonium nitrogen (NH4 + -N) is gradually oxidized to nitrate (NO3 - -N) is one of the key links in the nitrogen cycle in the natural environment. Nitrification is mainly completed by two types of nitrifying microorganisms: ammonia-oxidizing bacteria (AOB) and nitrite-oxidizing bacteria (NOB). Ammonia-oxidizing bacteria convert ammonium nitrogen (NH4 + -N) is oxidized to nitrite (NO2 - -N), and nitrite oxidizing bacteria further oxidize nitrite to nitrate. Nitrification is widely present in ecosystems such as soil, freshwater and ocean, and has an important impact on the ecological environment and agricultural production.

[0003] Excessive nitrification may bring about a series of environmental problems, such as groundwater pollution caused by nitrate leaching and reduced nitrogen fertilizer utilization. Therefore, how to effectively regulate the nitrification process to reduce adverse effects has become an important issue that needs to be solved in the fields of agriculture and environmental management. In recent years, research and development of new nitrification inhibitors to selectively inhibit the activity of ammonia oxidizing bacteria has become an important direction for achieving efficient nitrogen utilization and environmental protection. Summary of the invention

[0004] The present invention aims to develop a novel nitrification inhibitor to enrich the selection of nitrification inhibitors.

[0005] In order to solve the above problems, the present invention provides an application of 4-methyl-1-piperazineethylamine as a nitrification inhibitor and a method for inhibiting nitrification.

[0006] As a first aspect, the present invention relates to the use of 4-methyl-1-piperazineethylamine as a nitrification inhibitor.

[0007] As a second aspect, the present invention also relates to the use of 4-methyl-1-piperazineethylamine in inhibiting the nitrosation activity of ammonia oxidizing bacteria.

[0008] As a third aspect, the present invention also relates to the use of 4-methyl-1-piperazineethylamine in inhibiting the nitrosation activity of Nitrosomonas.

[0009] Optionally, the nitrosomonas includes Nitrosomonas sp. CZ-4 and / or Nitrosomonasnitrosa strain WH-1.

[0010] As a fourth aspect, the present invention also relates to the use of 4-methyl-1-piperazineethylamine in soil treatment and / or sewage treatment.

[0011] As a fifth aspect, the present invention also relates to a method for inhibiting nitrification, comprising: treating soil or water containing ammonium nitrogen and ammonia oxidizing bacteria with 4-methyl-1-piperazineethylamine.

[0012] Optionally, based on the volume of the soil or water body as 100%, the concentration of the 4-methyl-1-piperazineethylamine is not less than 0.01‰ w / v.

[0013] Optionally, based on the volume of the soil or water body being 100%, the concentration of the 4-methyl-1-piperazineethylamine is 1‰ w / v.

[0014] As a sixth aspect, the present invention also relates to a nitrification inhibitor, the component of which includes 4-methyl-1-piperazineethylamine.

[0015] As a seventh aspect, the present invention also relates to the use of the above-mentioned nitrification inhibitor in soil treatment and / or sewage treatment.

[0016] The beneficial effects of the present invention compared to the prior art are:

[0017] The present invention has found the nitrification inhibition activity of 4-methyl-1-piperazine ethylamine through a large number of studies and experiments. After testing, 4-methyl-1-piperazine ethylamine can inhibit the growth of ammonia oxidizing bacteria exemplified by Nitrosomonas at a lower dose (0.01‰w / v), thereby efficiently inhibiting nitrification in soil or water, reducing farmland nitrogen loss and N2O emissions in soil and water. And at a concentration of 1‰w / v, 4-methyl-1-piperazine ethylamine has an inhibition rate of nitrification of Nitrosomonas of more than 96%. In addition, the 4-methyl-1-piperazine ethylamine of the embodiment of the present invention does not have ecotoxicity and potential bioaccumulation, has less influence on other heterotrophic microorganisms in the soil, and has good environmental friendliness. BRIEF DESCRIPTION OF THE DRAWINGS

[0018] Figure 1 This is a graph showing changes in nitrite nitrogen concentrations in the control group and each experimental group on the 4th day in Example 1 of the present invention;

[0019] Figure 2 The graph of the change of nitrite nitrogen concentration in the control group and each experimental group in Example 2 of the present invention in the first 4 days;

[0020] Figure 3 The graph is a graph showing the absorbance value changes at 600 nm for the mixed solution of the experimental group and the control group in Example 3 of the present invention. DETAILED DESCRIPTION

[0021] In order to make the above-mentioned objects, features and advantages of the present invention more obvious and easy to understand, the specific embodiments of the present invention are described in detail below in conjunction with the accompanying drawings. Although certain embodiments of the present invention are shown in the accompanying drawings, it should be understood that the present invention can be implemented in various forms and should not be interpreted as being limited to the embodiments described herein. On the contrary, these embodiments are provided to provide a more thorough and complete understanding of the present invention. It should be understood that the drawings and embodiments of the present invention are only for exemplary purposes and are not intended to limit the scope of protection of the present invention.

[0022] Unless otherwise defined, all technical and scientific terms used in this application have the same meanings as those commonly understood by those skilled in the art to which this application belongs; the terms used in the specification of this application are only for the purpose of describing specific implementation methods and are not intended to limit this application;

[0023] The term "including" and its variations used in this article are open inclusions, that is, "including but not limited to"; the term "based on" is "based at least in part on"; the term "one embodiment" means "at least one embodiment"; the term "another embodiment" means "at least one other embodiment"; the term "some embodiments" means "at least some embodiments"; the term "optionally" means "optional embodiments". The relevant definitions of other terms will be given in the following description. It should be noted that the concepts of "first" and "second" mentioned in the present invention are used to distinguish different objects, rather than to describe a specific order or a primary and secondary relationship. In addition, the terms "first" and "second" are only used for descriptive purposes and cannot be understood as indicating or implying relative importance or implicitly indicating the number of technical features indicated. Therefore, the features defined as "first" and "second" may explicitly or implicitly include one or more of the features. In the description of this application, unless otherwise specified, the meaning of "multiple" is two or more.

[0024] In order to enrich the types of nitrification inhibitors, the embodiments of the present invention provide a use of 4-methyl-1-piperazineethylamine as a nitrification inhibitor and a method for inhibiting nitrification.

[0025] As a first aspect, the present invention relates to the use of 4-methyl-1-piperazineethylamine as a nitrification inhibitor, specifically, to the use of 4-methyl-1-piperazineethylamine in inhibiting the nitrosation activity of ammonia oxidizing bacteria, and more specifically, to the use of 4-methyl-1-piperazineethylamine in inhibiting the nitrosation activity of Nitrosomonas. Among them, Nitrosomonas may include Nitrosomonas sp.CZ-4 (preservation number is CCTCCNO: M 2018813) and / or Nitrosomonas nitrosastrain WH-1 (preservation number is CCTCCNO: M 2020526). Further, based on the activity of 4-methyl-1-piperazineethylamine in inhibiting nitrification, the present invention also relates to the use of 4-methyl-1-piperazineethylamine in soil treatment and / or sewage treatment.

[0026] The present invention also specifically relates to a method for inhibiting nitrification, comprising: treating soil or water containing ammonium nitrogen and ammonia oxidizing bacteria with 4-methyl-1-piperazineethylamine. Specifically, based on 100% volume of the soil or water, the concentration of 4-methyl-1-piperazineethylamine is not less than 0.01‰w / v, and further, the concentration of 4-methyl-1-piperazineethylamine is preferably 1‰w / v.

[0027] The present invention also relates to a nitrification inhibitor, wherein the components of the nitrification inhibitor include 4-methyl-1-piperazineethylamine. Specifically, 4-methyl-1-piperazineethylamine can be used as the active component of the nitrification inhibitor. Based on the nitrification inhibition activity of the nitrification inhibitor, the present invention also relates to the use of the nitrification inhibitor in soil treatment and / or sewage treatment.

[0028] The present invention has found the nitrification inhibition activity of 4-methyl-1-piperazine ethylamine through a large number of studies and experiments. After testing, 4-methyl-1-piperazine ethylamine can inhibit the growth of ammonia oxidizing bacteria exemplified by Nitrosomonas at a lower dose (0.01‰w / v), thereby efficiently inhibiting nitrification in soil or water, reducing farmland nitrogen loss and N2O emissions in soil and water. And at a concentration of 1‰w / v, 4-methyl-1-piperazine ethylamine has an inhibition rate of nitrification of Nitrosomonas of more than 96%. In addition, the 4-methyl-1-piperazine ethylamine of the embodiment of the present invention does not have ecotoxicity and potential bioaccumulation, has less influence on other heterotrophic microorganisms in the soil, and has good environmental friendliness.

[0029] The present invention is described in detail below with reference to specific embodiments and comparative examples:

[0030] It should be noted that the solutions and preparation methods used in the following examples are as follows:

[0031] Trace element solution: includes the following components in mass concentrations: 0.5 g / L EDTA, 0.075 g / L CuSO4·5H2O, 0.3 g / L ZnSO4·7H2O, 0.375 g / L CoCl2·6H2O, 0.3 g / L MnCl2·2H2O, 0.014 g / L H3BO4 and 0.22 g / L NaMoO4·2H2O.

[0032] Ammonia oxidizing bacteria medium (AOB): FeSO4·7H2O, MgSO4·7H2O, K2HPO4, NaHCO3, CaCl2·2H2O, NaCl, NH4Cl and trace element solution were dissolved in water. The mass concentration of FeSO4·7H2O, MgSO4·7H2O, K2HPO4, NaHCO3, CaCl2·2H2O, NaCl, NH4Cl and trace element solution were controlled to be 0.4 g / L, 0.5 g / L, 0.1 g / L, 1.5 g / L, 0.5 g / L, 2.0 g / L, 0.4 g / L and 1 mL / L of trace element aqueous solution. The pH of the ammonia oxidizing bacteria medium was controlled to be 7.5, and then sterilized under high temperature and high pressure (121°C, 20 min) and stored for later use.

[0033] Beef peptone medium (BPM): purchased from Qingdao Haibo Biological, prepared at a concentration of 18 g / L, sterilized at high temperature and high pressure (121°C, 20 min) and stored for future use.

[0034] 4-Methyl-1-piperazineethylamine mother liquor: 1 g of 4-methyl-1-piperazineethylamine (CAS: 934-98-5, purchased from McLean) was weighed using an analytical balance, dissolved with a small amount of water, and then transferred to a 100 mL volumetric flask to obtain a 4-methyl-1-piperazineethylamine mother liquor. The 4-methyl-1-piperazineethylamine mother liquor was then filtered and sterilized using a 0.22 μm filter membrane and stored for later use.

[0035] It should be further explained that in the embodiments of the present invention, the ammonia nitrogen concentration detection method adopts the "Water Quality - Determination of Ammonia Nitrogen - Nessler's Reagent Spectrophotometric Method" (HJ 535-2009); the nitrite nitrogen concentration is determined by the "Water Quality - Determination of Nitrite Nitrogen - Spectrophotometric Method" (GB / T 7493-1987).

[0036] It should be further explained that in the embodiment of the present invention, the nitrification inhibition rate is calculated using the following formula:

[0037]

[0038] Example 1

[0039] This example relates to an experiment to evaluate the nitrification inhibition effect of 4-methyl-1-piperazineethylamine on the bacterial species Nitrosomonas sp. CZ-4 (hereinafter referred to as CZ-4).

[0040] Take 1 mL of CZ-4 bacterial liquid into fresh AOB medium and culture it in a shaking incubator at 30°C and 140 r / min. Take samples every 24 hours to test the ammonia nitrogen concentration and nitrite nitrogen concentration until the growth rate of nitrite nitrogen concentration reaches 2 mg / L / h.

[0041] The 4-methyl-1-piperazineethylamine mother solution was added to the AOB medium containing the CZ-4 bacterial solution at different concentrations, and the CZ-4 culture system was prepared according to the formula in Table 1. Among them, the CZ-4 culture system without the addition of the 4-methyl-1-piperazineethylamine mother solution was used as the control group, and the other CZ-4 culture systems were used as the experimental group. Then, the culture was carried out in a shaking table at 30°C and 140r / min.

[0042] Table 1 CZ-4 experimental system formula for the control group and experimental group in Example 1

[0043]

[0044] After 4 days, samples were taken to measure the concentrations of ammonia nitrogen and nitrite nitrogen in the control group and each experimental group. The concentrations of nitrite nitrogen in each control group and each experimental group on the 4th day are shown in Table 2 and Figure 1 As shown:

[0045] Table 2 Nitrite nitrogen concentrations of the control groups and experimental groups in Example 1

[0046]

[0047] From Table 2 and Figure 1 It can be seen that the nitrite nitrogen concentration of the control group reached 80.76 mg / L on the 4th day, indicating that the bacteria grew well. After the addition of 4-methyl-1-piperazineethylamine, the increase in nitrite nitrogen concentration in the culture system was inhibited to varying degrees, indicating that the growth of CZ-4 was inhibited to varying degrees. And as the concentration decreased, the inhibition ability decreased, and there was no inhibition at a concentration of 0.001‰w / v; at a concentration of 0.01‰w / v, the nitrification inhibition rate was 3.38%; at a concentration of 1‰w / v, the nitrification inhibition rate reached 96.87%.

[0048] Example 2

[0049] This example relates to an experiment to evaluate the nitrification inhibition effect of 4-methyl-1-piperazineethylamine on the bacterial strain Nitrosomonas nitrosa WH-1 (hereinafter referred to as WH-1).

[0050] Take 1 mL of WH-1 bacterial solution and add it to fresh AOB medium. Culture it in a shaking incubator at 30°C and 140 r / min. Take samples every 24 hours to test the ammonia nitrogen concentration and nitrite nitrogen concentration until the growth rate of nitrite nitrogen concentration reaches 2 mg / L / h.

[0051] The 4-methyl-1-piperazineethylamine mother solution was added to the AOB medium containing the WH-1 bacterial solution at different concentrations, and the WH-1 culture system was prepared according to the formula in Table 3. Among them, the WH-1 culture system without the addition of the 4-methyl-1-piperazineethylamine mother solution was used as the control group, and the other WH-1 culture systems were used as the experimental group. Then, the shaking table was cultured at 30°C and 140r / min, and the concentrations of ammonia nitrogen and nitrite nitrogen were measured every day.

[0052] Table 3 WH-1 experimental system formula for the control group and experimental group in Example 2

[0053]

[0054] The nitrite nitrogen concentrations of the control group and the experimental group on d1, d2, d3 and d4 are shown in Table 4 and Figure 2 As shown:

[0055] Table 4 Nitrite nitrogen concentrations in each control group and each experimental group in Example 2

[0056]

[0057]

[0058] From Table 4 and Figure 2 It can be seen that the nitrite nitrogen concentration in the control group rose sharply after 3 days, tended to be flat on the 4th day, and entered a stable period. After adding 4-methyl-1-piperazineethylamine, the increase of nitrite nitrogen in the WH-1 culture system was inhibited to varying degrees, indicating that the growth of WH-1 was inhibited to varying degrees, and the inhibition ability decreased with the decrease of concentration.

[0059] From Table 4 and Figure 2 It can be further known that at a concentration of 0.001‰w / v of 4-methyl-1-piperazineethylamine, the WH-1 experimental system was almost not inhibited, and at a concentration of 0.01‰w / v of 4-methyl-1-piperazineethylamine, the WH-1 experimental system was weakly inhibited, and the nitrification inhibition rate was 2.43%; at a concentration of 0.1‰w / v of 4-methyl-1-piperazineethylamine, the nitrification inhibition rate of the WH-1 experimental system was 9.47%, and the inhibition began to show obvious effects on the third day; at a concentration of 1‰w / v of 4-methyl-1-piperazineethylamine, the nitrification inhibition rate of the WH-1 experimental system reached 96.85%, and the inhibition began to show obvious effects on the second day.

[0060] Example 3

[0061] This example relates to an experiment to evaluate the inhibition of 4-methyl-1-piperazineethylamine on heterotrophic microorganisms in soil.

[0062] Specifically, 20 grams of soil and 20 glass beads were added to 200 milliliters of normal saline, shaken at a speed of 180r / min for 30 minutes, and then settled for 5 minutes, and the supernatant was collected as the soil microbial inoculum. Two groups of 20mL BPM culture medium were placed in sterile conical flasks respectively, and 20μL of soil microorganisms were inoculated into the two groups of BPM culture medium respectively. One group was then added with 4-methyl-1-piperazineethylamine at a concentration of 0.01‰w / v as the experimental group; the other group was not treated as the control group. After the experimental group and the control group were cultured in a shaking incubator at 30°C and 140r / min overnight, the absorbance of the mixed solution of the experimental group and the control group at 600nm was measured using a visible spectrophotometer. The results are as follows Figure 3 shown.

[0063] Depend on Figure 3 It can be seen that after 16 hours, the absorbance values ​​of the mixed solution of the control group and the experimental group at 600nm were 1.5 and 1.4 respectively. This shows that after adding 0.01‰w / v of 4-methyl-1-piperazineethylamine, the growth of microorganisms in the soil extract is relatively consistent with that of the blank control group, indicating that 0.01‰w / v of 4-methyl-1-piperazineethylamine has little effect on other heterotrophic microorganisms in the soil and has good environmental friendliness.

[0064] Although the present invention is disclosed as above, the protection scope of the present invention is not limited thereto. Those skilled in the art may make various changes and modifications without departing from the spirit and scope of the present invention, and these changes and modifications will fall within the protection scope of the present invention.

Claims

Application of 1.4-methyl-1-piperazineethylamine as a nitrification inhibitor. Application of 2.4-methyl-1-piperazineethylamine in inhibiting the nitrosation activity of ammonia oxidizing bacteria.

3. Application of 4-methyl-1-piperazineethylamine in inhibiting the nitrosation activity of Nitrosomonas.

4. The use according to claim 3, characterized in that: The nitrosomonas include Nitrosomonas sp. CZ-4 and / or Nitrosomonas nitrosa strain WH-1.

5. Application of 4-methyl-1-piperazineethylamine in soil treatment and / or sewage treatment.

6. A method for inhibiting nitrification, characterized in that: include: Use 4-methyl-1-piperazineethylamine to treat soil or water containing ammonium nitrogen and ammonia oxidizing bacteria.

7. The method for inhibiting nitrification according to claim 6, characterized in that: Based on the volume of the soil or water body as 100%, the concentration of the 4-methyl-1-piperazineethylamine is not less than 0.01‰ w / v.

8. The method for inhibiting nitrification according to claim 6, characterized in that: Based on the volume of the soil or water body being 100%, the concentration of the 4-methyl-1-piperazineethylamine is 1‰ w / v.

9. A nitrification inhibitor, characterized in that: The nitrification inhibitor component includes 4-methyl-1-piperazineethylamine.

10. Use of the nitrification inhibitor according to claim 9 in soil treatment and / or sewage treatment.

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