Application of phospholipase in promoting removal of phosphorus in water body

Abstract

The invention discloses application of phospholipase in promoting removal of phosphorus in a water body, and belongs to the technical field of water treatment. According to the method, phospholipase is added into a wetland or artificial wetland system, and the phospholipase catalyzes and degrades organic phosphorus into an intermediate, so that the time for converting the organic phosphorus into absorbable phosphorus is shortened, the conversion of phosphorus into absorbable phosphorus is accelerated, and the phosphorus absorption efficiency of plants is improved, thereby improving the removal efficiency of the wetland or artificial wetland system on phosphorus in a water body.

Description

Technical Field

[0001] The present application belongs to the field of water treatment technology, and specifically relates to the application of phospholipase in promoting the removal of phosphorus in water. Background Art

[0002] The phosphorus load generated by human production and life has led to eutrophication of lakes. Therefore, in the deep treatment of sewage, phosphorus removal is one of the key links in water management.

[0003] Wetlands or artificial wetlands, including unique soil (filler)-plant-microorganism ecosystems, are cheap and effective application technologies for controlling phosphorus loads. The mechanisms for removing phosphorus from water bodies are mainly physical, chemical and biological. Physical effects include deposition, which mainly refers to the process in which soluble phosphates in the influent are physically stored in the wetland; chemical effects include adsorption and precipitation, which mainly refer to the adsorption of phosphorus by soil (filler) and precipitation in wetlands or artificial wetland systems; biological effects include plant absorption and normal absorption and excessive accumulation of microorganisms. Plant absorption mainly refers to inorganic phosphate (PO4 3- 、HPO4 2- 、H2PO4 - ) is absorbed by plant roots and assimilated into the plant's organic components (such as ATP, phospholipids, coenzymes, DNA and RNA, etc.), and then taken out of the system through harvesting. Normal absorption and excessive accumulation of microorganisms mainly refer to the absorption of phosphorus into cells during microbial growth and its conversion into components of microbial cells.

[0004] Among them, chemical action is considered to be one of the most important mechanisms for phosphorus removal, so improving the adsorption capacity of soil (filler) has become a hot topic of research. For example, the Chinese invention patent with publication number CN105776552A records a phosphorus removal filler for artificial wetlands and its preparation method, providing a core-shell structure phosphorus removal filler with a core composed of Ca(OH)2 and trivalent iron salt and an outer layer covered with CaCO3; the Chinese invention patent with publication number CN112979086A records an enhanced artificial wetland phosphorus removal filler, which is composed of a composite filler and a polyphosphate bacteria activation liquid, and the composite filler is selected from zeolite, medical stone, crushed stone, magnetite, volcanic stone, dolomite, shale ceramsite, oyster shell, sea Any combination of clam shells, coal slag, slag, peat, coarse sand, gravel, clay ceramsite, fly ash, and steel slag; Chinese invention patent with publication number CN115180713A records a high-efficiency phosphorus removal wetland filler and its preparation method, and the high-efficiency phosphorus removal wetland filler includes: gravel, soil, shale, limestone, manganese sand, sludge, zeolite and fly ash, wherein the gravel accounts for 40-60%, soil 10-15%, shale 5-8%, limestone 5-8%, manganese sand 3-5%, sludge 10-15%, zeolite 2-3% and fly ash 4-6%. However, after the system has been running for a period of time, the phosphorus removal rate of the soil (filler) will decrease to a certain extent, and the adsorbed and precipitated phosphorus still exists in the wetland or artificial wetland system, and there is still a risk of cyclic release.

[0005] Correspondingly, plant absorption deserves more attention because it absorbs phosphorus and assimilates it into organic components of plants and is then taken out of the system through harvesting. However, it also has certain limitations, mainly because phosphorus in sewage is often present in the form of inorganic phosphate (PO4 3- 、HPO4 2- 、H2PO4 - ), polyphosphates and organic phosphorus, among which only inorganic phosphates can be absorbed by plants. Although organic phosphorus can be gradually converted into inorganic phosphate (i.e. absorbable phosphorus) absorbable by plants through a variety of complex pathways under natural conditions, its conversion process is very slow and time-consuming. Often, the sewage is discharged from the system before the conversion process is completed, which affects the removal efficiency of phosphorus in the water, is not conducive to improving the effluent water quality, and increases the risk of eutrophication of the receiving water body. Summary of the invention

[0006] 1. Problem to be solved

[0007] The present application aims to solve the problem that plants in wetland or artificial wetland systems are unable to absorb organic phosphorus, resulting in low phosphorus absorption efficiency of plants. The application of phospholipase in promoting the removal of phosphorus in water bodies is provided. Through the catalytic action of phospholipase, organic phosphorus is converted into intermediates, thereby shortening the time for converting organic phosphorus to absorbable phosphorus, accelerating the conversion of phosphorus to absorbable phosphorus, improving the phosphorus absorption efficiency of plants, and promoting the removal efficiency of phosphorus in water bodies.

[0008] 2. Technical solution

[0009] In order to solve the above problems, the technical solutions adopted in this application are as follows:

[0010] The present application provides the use of phospholipase in promoting the removal of phosphorus from water bodies. The application is to add phospholipase to a wetland or artificial wetland system, and the phospholipase catalyzes the degradation of organic phosphorus into intermediates, thereby shortening the time for organic phosphorus to be converted into absorbable phosphorus, accelerating the conversion of phosphorus into absorbable phosphorus, and improving the absorption efficiency of phosphorus by plants, thereby improving the removal efficiency of phosphorus from water bodies by the wetland or artificial wetland system; phospholipase (phospholipases) is a class of enzymes responsible for phospholipid metabolism and biosynthesis in organisms, and can catalyze the hydrolysis reaction of glycerophospholipids; phospholipase is generally widely used in the field of food processing, and the purpose of hydrolyzing phospholipids is to improve the taste of food, such as improving the flavor of pasta, or focusing on the role in plant phosphorus circulation, such as in the field of crops, and is basically not involved in environmental protection water treatment.

[0011] Furthermore, the phospholipase includes any one or more of phospholipase A1 (PLA1), phospholipase A2 (PLA2), phospholipase B (PLB), phospholipase C (PLC) and phospholipase D (PLD); the phospholipase is classified according to the different phospholipid hydrolysis sites, and the phospholipid hydrolysis sites are as follows: Figure 1As shown in the figure, phospholipase A1 (PLA1) and phospholipase A2 (PLA2) mainly act on specific ester bonds in phospholipid molecules (such as the 1st or 2nd ester bonds of glycerophospholipids), hydrolyzing to produce fatty acids (such as arachidonic acid) and lysophospholipids, and do not directly hydrolyze phosphate; phospholipase B (PLB) has a wider substrate specificity and can hydrolyze the two ester bonds of phospholipids, including those containing phosphate groups, but its main product is not single phosphate, but free fatty acids and glycerophosphatidylcholine; phospholipase C (PLC) mainly acts on The phospholipids have phosphate bonds, but their main products are diacylglycerol and phosphorylated head groups, not separate phosphate groups; phospholipase D (PLD) mainly acts on the head groups of phospholipids to produce phosphatidic acid and alcohols, and also does not directly produce phosphate groups; however, although the products of the above-mentioned phospholipid hydrolysis are not inorganic phosphates (i.e., absorbable phosphorus) that can be directly absorbed by plants, the present invention has found that after being treated with phospholipase, the absorption efficiency of phosphorus by plants can be promoted. It is speculated that the conversion of organic phosphorus into absorbable phosphorus is accelerated after the organic phosphorus is converted into intermediates.

[0012] Furthermore, the above-mentioned phospholipase includes phospholipase A1 and / or phospholipase B.

[0013] Furthermore, the above application includes: using nanospheres as filler matrix, loading the above phospholipase, and then wrapping with sodium alginate to use as artificial wetland filler; after the phospholipase is loaded and wrapped, the phospholipase can be fixed and wrapped, so that the phospholipase is slowly released, the effective time of the phospholipase is extended, and the lasting effectiveness of the phospholipase on the flowing water body is achieved.

[0014] Furthermore, the above artificial wetland filler also includes CaCl2, Ca 2+ It plays a role in promoting phospholipase hydrolysis. Metal ions such as calcium ions can promote the hydrolysis reaction by interacting with specific groups in organic phosphorus molecules.

[0015] Furthermore, the preparation method of the above-mentioned artificial wetland filler includes: adding nanospheres to a phospholipase solution, shaking and then standing to obtain catalytic nanospheres; adding the catalytic nanospheres to a sodium alginate solution, stirring until the surface is covered with sodium alginate; then putting them into a calcium chloride solution, embedding and taking them out to obtain the above-mentioned artificial wetland filler.

[0016] Furthermore, the above-mentioned artificial wetland filler also includes indoleacetic acid, vitamin B1, vitamin B6, calcium, iron, zinc, magnesium, etc.; it can promote the growth of plant roots, thereby improving the absorption efficiency of phosphorus.

[0017] Furthermore, the preparation method of the above-mentioned artificial wetland filler includes: adding nanospheres to a mixed auxiliary liquid containing indoleacetic acid, vitamin B1, vitamin B6, calcium, iron, zinc and magnesium, shaking and then standing to obtain auxiliary nanospheres; adding catalytic nanospheres and auxiliary nanospheres to a sodium alginate solution, stirring until their surfaces are covered with sodium alginate; then putting them into a calcium chloride solution, embedding and taking them out to obtain the above-mentioned artificial wetland filler.

[0018] Furthermore, the above calcium is calcium chloride.

[0019] Furthermore, the above-mentioned iron is ferrous sulfate.

[0020] Furthermore, the above zinc is zinc sulfate.

[0021] Furthermore, the magnesium is magnesium sulfate. Furthermore, in the mixed auxiliary liquid, the concentration of indoleacetic acid is 10-1000 mg / L. Furthermore, in the mixed auxiliary liquid, the concentration of indoleacetic acid is 1000 mg / L.

[0022] Furthermore, in the above-mentioned mixed auxiliary liquid, the concentration of vitamin B1 is 1-100 mg / L. Furthermore, in the above-mentioned mixed auxiliary liquid, the concentration of vitamin B1 is 100 mg / L.

[0023] Furthermore, in the above-mentioned mixed auxiliary liquid, the concentration of vitamin B6 is: 1-100 mg / L. Furthermore, in the above-mentioned mixed auxiliary liquid, the concentration of vitamin B6 is 100 mg / L.

[0024] Furthermore, in the above-mentioned mixed auxiliary liquid, the concentration of calcium chloride is 10-10000 mg / L. Furthermore, in the above-mentioned mixed auxiliary liquid, the concentration of calcium chloride is 10000 mg / L.

[0025] Furthermore, in the above-mentioned mixed auxiliary liquid, the concentration of ferrous sulfate is 10-10000 mg / L. Furthermore, in the above-mentioned mixed auxiliary liquid, the concentration of ferrous sulfate is 10000 mg / L.

[0026] Furthermore, in the above-mentioned mixed auxiliary liquid, the concentration of zinc sulfate is 1-1000 mg / L. Furthermore, in the above-mentioned mixed auxiliary liquid, the concentration of zinc sulfate is 1000 mg / L.

[0027] Furthermore, in the above-mentioned mixed auxiliary liquid, the concentration of magnesium sulfate is 0.5-500 mg / L. Furthermore, in the above-mentioned mixed auxiliary liquid, the concentration of magnesium sulfate is 500 mg / L.

[0028] Furthermore, the mass ratio of the nanospheres to the phospholipase solution is 1:2 or above, and the phospholipase solution can completely immerse the nanospheres.

[0029] Furthermore, the mass ratio of the nanospheres to the mixed auxiliary liquid is 1:2 or above, and the mixed auxiliary liquid can completely immerse the nanospheres.

[0030] Furthermore, the mass ratio of the catalytic nanospheres to the auxiliary nanospheres is 9:1 or above.

[0031] Furthermore, the sodium alginate solution is 4% sodium alginate, which is prepared by adding 4 grams of sodium alginate powder to every 100 milliliters of distilled water, stirring thoroughly until uniform and transparent, and then standing for 2 to 3 hours.

[0032] Furthermore, the calcium chloride solution is 4% calcium chloride, which is prepared by adding 4 grams of calcium chloride to every 100 milliliters of distilled water and stirring thoroughly until it is uniform and transparent.

[0033] The present application also provides an artificial wetland filler for promoting the removal of phosphorus from water bodies, the filler comprising:

[0034] Nanospheres,

[0035] Phospholipase, including any one or more of phospholipase A1 (PLA1), phospholipase A2 (PLA2), phospholipase B (PLB), phospholipase C (PLC) and phospholipase D (PLD),

[0036] Sodium alginate,

[0037] Calcium chloride,

[0038] The phospholipase is loaded on the nanospheres, and the sodium alginate wraps the nanospheres loaded with the phospholipase.

[0039] Furthermore, the above-mentioned artificial wetland filler also includes: indoleacetic acid, vitamin B1, vitamin B6, calcium, iron, zinc, and magnesium.

[0040] The present application also provides an artificial wetland system, in which the filler includes the above-mentioned artificial wetland filler for promoting the removal of phosphorus from water bodies.

[0041] The present application also provides the above-mentioned artificial wetland filler for promoting the removal of phosphorus from water bodies and the use of the above-mentioned artificial wetland system in the removal of phosphorus from water bodies.

[0042] 3. Beneficial effects

[0043] Compared with the prior art, the present application has the following beneficial effects:

[0044] (1) The present application provides an application of phospholipase in promoting the removal of phosphorus from water bodies. Although phospholipase cannot directly convert organic phosphorus into absorbable phosphorus, the inventors unexpectedly discovered and verified through experiments that the intermediate product after phospholipase-catalyzed hydrolysis can shorten the time for converting organic phosphorus into absorbable phosphorus, thereby accelerating the conversion of phosphorus into absorbable phosphorus. In other words, the use of phospholipase promotes the conversion of organic phosphorus into absorbable phosphorus, improves the efficiency of plant absorption of phosphorus, and thereby improves the removal efficiency of phosphorus in wetland or artificial wetland systems.

[0045] (2) The application of the phospholipase provided in the present application in promoting the removal of phosphorus from water bodies is to use nanospheres as filler matrix, load the phospholipase and then encapsulate it with sodium alginate, and then use it as artificial wetland filler to fix and encapsulate the phospholipase, so that the phospholipase is slowly released, prolonging the effective time of the phospholipase, and realizing the long-term effectiveness of the phospholipase on flowing water bodies.

[0046] (3) The use of the phospholipase provided in the present application in promoting the removal of phosphorus from water, wherein the filler containing the phospholipase also includes CaCl2, indoleacetic acid, vitamin B1, vitamin B6, calcium, iron, zinc, magnesium, etc.; wherein Ca 2+ It plays a role in promoting phospholipase hydrolysis. Indoleacetic acid, vitamin B1, vitamin B6, calcium, iron, zinc, magnesium, etc. can promote the growth of plant roots, thereby improving the adsorption efficiency of phosphorus. BRIEF DESCRIPTION OF THE DRAWINGS

[0047] Figure 1 This is a map of the sites where different phospholipases hydrolyze phospholipids.

[0048] Figure 2 This is the simulated artificial wetland system of Example 2.

[0049] Figure 3 It is the change of organic phosphorus concentration in the simulated artificial wetland system of Example 2.

[0050] Figure 4 It is the change of total phosphorus concentration in the simulated artificial wetland system of Example 2.

[0051] Figure 5 It is the cumulative removal rate of total phosphorus in the simulated artificial wetland system of Example 2.

[0052] Figure 6 This is the simulated artificial wetland system of Example 3.

[0053] Figure 7 It is the change of organic phosphorus concentration in the simulated artificial wetland system of Example 3.

[0054] Figure 8 It is the change of total phosphorus concentration in the simulated artificial wetland system of Example 3.

[0055] Fig. 9 It is the daily removal rate of total phosphorus in the simulated artificial wetland system of Example 3.

[0056] Fig.10 It is the cumulative removal rate of total phosphorus in the simulated artificial wetland system of Example 3.

[0057] Fig.11 It is the height change of water hyacinth in the simulated artificial wetland system of Example 3.

[0058] Fig.12 It is the height change of Acorus calamus in the simulated artificial wetland system of Example 3.

[0059] Fig.13 It is the height change of Acorus calamus in the simulated artificial wetland system of Example 3. DETAILED DESCRIPTION

[0060] The present application is further described below in conjunction with specific embodiments.

[0061] It should be noted that the terms such as "upper", "lower", "left", "right", "middle", etc. cited in this specification are only for the convenience of description and are not used to limit the scope of implementation. Changes or adjustments to their relative relationships should be regarded as the scope of implementation of this application without substantially changing the technical content.

[0062] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by those skilled in the art to which this application belongs; the term "and / or" used herein includes any and all combinations of one or more of the associated listed items.

[0063] If the specific conditions are not specified in the examples, the experiments were carried out under conventional conditions or conditions recommended by the manufacturer. If the manufacturers of the reagents or instruments are not specified, they are all conventional products that can be purchased commercially.

[0064] As used herein, the term "about" is used to provide flexibility and imprecision associated with a given term, measurement or value. The degree of flexibility for a particular variable can be easily determined by one skilled in the art.

[0065] As used herein, the term "at least one of" is intended to be synonymous with "one or more of." For example, "at least one of A, B, and C" expressly includes only A, only B, only C, and combinations of each thereof.

[0066] Concentration, amount and other numerical data can be presented in range format herein.It should be understood that such range format is only used for convenience and simplicity, and should be flexibly interpreted as not only including the numerical value clearly described as range limit, but also including all single numerical values ​​or sub-ranges contained in the range, just as each numerical value and sub-range are clearly described.For example, the numerical range of about 1 to about 4.5 should be interpreted as not only including the limit value of 1 to about 4.5 clearly described, but also including single numerals (such as 2,3,4) and sub-ranges (such as 1 to 3,2 to 4, etc.).The same principle is applicable to the scope of only narrating a numerical value, such as "less than about 4.5", which should be interpreted as including all the above-mentioned values ​​and ranges.In addition, no matter how the breadth of the described range or feature is, this explanation should be applicable.

[0067] Those skilled in the art know that there are many kinds of plants in artificial wetland systems, which can be selected according to factors such as location, climate, and water samples. In this application, the plants used to simulate artificial wetland systems include a combination of two or three of water hyacinth (origin is Yongzhou, Hunan), water calamus (origin is Suqian, Jiangsu), and stone calamus (origin is Lishui, Zhejiang), and the above plants can be obtained commercially; before the experiment, the plants are pre-cultured according to their growth characteristics and experimental requirements, and the culture time is 5 to 7 days to allow the plants to adapt to the laboratory environment.

[0068] In the present application, phospholipase A1 was purchased from Novozymes (Denmark, novozymes, Lecitase Ultra, 10 KLU / g, 50 g), and phospholipase B was purchased from Cangzhou Xiasheng Enzyme Biotechnology Co., Ltd. (Cat. No.: FDG-2238, 0.1 kg, 2900 U / g enzyme activity).

[0069] In the present application, soybean lecithin was used to simulate the organic phosphorus in water. The soybean lecithin was purchased from Henan Jiapai Biotechnology Co., Ltd., 100 g / bag.

[0070] Example 1

[0071] This embodiment provides the application of phospholipase in promoting the removal of phosphorus from water bodies, specifically using nanospheres as filler matrix, loading phospholipase and then wrapping with sodium alginate to use as artificial wetland filler.

[0072] The artificial wetland filler includes:

[0073] Nanospheres, as filler matrix for constructed wetland fillers, are used to load phospholipase;

[0074] Phospholipases, as catalytically active components, specifically break specific chemical bonds in organophosphates;

[0075] Indoleacetic acid, vitamin B1, vitamin B6, calcium, iron, zinc, magnesium, used to promote plant root growth; Ca 2+ It can also promote the hydrolysis reaction of phospholipase;

[0076] Sodium alginate is used to fix and encapsulate phospholipase and indoleacetic acid, vitamin B1, vitamin B6, calcium, iron, zinc and magnesium.

[0077] The preparation method of the artificial wetland filler comprises:

[0078] (1) Solution preparation

[0079] The nanospheres (nanobacteria spheres, Maibao) were washed with distilled water and dried at 103-105°C for later use;

[0080] Prepare a mixed auxiliary solution: add 1g indoleacetic acid, 0.1g vitamin B1, 0.1g vitamin B6, 10g calcium chloride, 10g ferrous sulfate, 1g zinc sulfate, and 0.5g magnesium sulfate to 1 liter of water;

[0081] Prepare 4% sodium alginate solution: add 4 g of sodium alginate powder to every 100 ml of distilled water, stir thoroughly until uniform and transparent, and let stand for 2 to 3 hours;

[0082] Prepare 4% calcium chloride solution: add 4 g of calcium chloride to every 100 ml of distilled water and stir thoroughly until it is homogeneous and transparent;

[0083] (2) Preparation

[0084] The nanospheres are added to the phospholipase solution at a mass ratio of 1:2, and the solution is shaken at 20-30° C. for more than 30 minutes, and then allowed to stand for 24 hours to obtain catalytic nanospheres;

[0085] The nanospheres were added into the mixed auxiliary solution, the mass ratio of the nanospheres to the phospholipase solution was 1:2, the mixture was shaken at 20-30°C for 1 hour, and allowed to stand for more than 2 hours to obtain auxiliary nanospheres;

[0086] Add catalytic nanospheres and auxiliary nanospheres into sodium alginate solution at a mass ratio of 9:1, and stir until the surface is covered with sodium alginate; then put them into calcium chloride solution, embed and take them out to obtain artificial wetland filler.

[0087] In this embodiment, the phospholipase solution is 10KLU / g phospholipase A1, and an artificial wetland filler containing phospholipase A1 is prepared; the adsorption amount of the nanospheres to the solution is 10% on average. When the concentration of the phospholipase A1 solution is 10KLU / g, the catalytic nanospheres prepared from 10g of nanospheres have a mass of about 11g, corresponding to 10KLU phospholipase A1; the catalytic nanospheres and the auxiliary nanospheres are in a mass ratio of 9:1, and the mass of the auxiliary nanospheres is about 1.2g; about 12.2g of filler corresponds to 10KLU phospholipase A1.

[0088] In this embodiment, the phospholipase solution is 10g / L phospholipase B, and an artificial wetland filler containing phospholipase B is prepared; the adsorption amount of the nanospheres to the solution is 10% on average. When the concentration of the phospholipase B solution is 10g / L, the catalytic nanospheres prepared from 20g of nanospheres have a mass of about 22g and adsorb about 2mL by volume (the density of the solution is calculated based on the approximate water), which corresponds to about 20mg of phospholipase B; because the mass ratio of the catalytic nanospheres and the auxiliary nanospheres is 9:1, the mass of the auxiliary nanospheres is about 2.4g; about 24.4g of filler corresponds to 20mg of phospholipase B.

[0089] Example 2

[0090] This example provides a study on the effects of different types of phospholipases on phosphorus removal in water.

[0091] In this embodiment, the plants used to simulate the artificial wetland system include water hyacinth and calamus. Figure 2 As shown, water hyacinth (9 to 10 pieces, about 5 cm in height) and calamus (3 to 4 pieces, about 30 cm in height).

[0092] In this embodiment, the experiment was divided into four groups (including the control group), and 2L of natural lake water (water quality was better than Class III water in the "Surface Water Environmental Quality Standard") was added to each group of simulated artificial wetland systems, and 10mL of 25mg / mL soybean lecithin emulsion was added to simulate wastewater containing organic phosphorus; the types and amounts of artificial wetland fillers prepared by phospholipase in the four groups of experiments were as follows:

[0093] (1) Phospholipase B artificial wetland filler experimental group: 73.2 g of the artificial wetland filler containing phospholipase B prepared in Example 1 was added (i.e., about 60 mg of phospholipase B was added);

[0094] (2) Phospholipase A1 artificial wetland filler experimental group: 12.2 g of the artificial wetland filler containing phospholipase A1 prepared in Example 1 was added (i.e., about 10 KLU of phospholipase A1 was added);

[0095] (3) Experimental group of combination of phospholipase A1 artificial wetland filler and phospholipase B artificial wetland filler: 73.2 g of artificial wetland filler containing phospholipase B prepared in Example 1 and 12.2 g of artificial wetland filler containing phospholipase A1 prepared in Example 1 were added;

[0096] (4) No enzyme experimental group: no phospholipase was added;

[0097] The experiment will last from April 14, 2024 to April 28, 2024, a total of 15 days.

[0098] Result analysis:

[0099] (1) Effects of different types of phospholipases on organic phosphorus removal

[0100] The concentration changes of organic phosphorus in the simulated artificial wetland system are as follows Figure 3 As shown, the organic phosphorus concentration and removal rate are shown in Table 1. It can be seen from Table 1 that within one day of adding artificial wetland fillers with phospholipase B, phospholipase A1, and a combination of phospholipase A1 and phospholipase B, the concentration of organic phosphorus in the water samples of the simulated artificial wetland system decreased by 79.9%, 85.8% and 89.6%, respectively, while the concentration of organic phosphorus in the blank sample decreased by 12.1%. This shows that the effect of phospholipase A1 in promoting the conversion of lecithin into soluble orthophosphate is slightly stronger than that of phospholipase B within 24 hours, but within 48 hours and longer time ranges (such as 1 to 2 weeks), phospholipase B has a stronger removal effect. And the combination of the two has a better effect on the removal of organic phosphorus in water. The reason for this is that phospholipase B and phospholipase A1 can hydrolyze lecithin into lysophospholipids and a free fatty acid, and can further hydrolyze the phosphate bonds in lysophospholipids, thereby further releasing phosphate groups, so that organic phosphorus can be removed. Figure 3 It can be seen that the concentration of organic phosphorus in the three simulated artificial wetland systems with phospholipase first dropped rapidly, then stabilized and showed a fluctuating state; the concentration of organic phosphorus in the blank sample first dropped steadily, then stabilized and showed a fluctuating state. By comparing the three, the following conclusions can be drawn: Phospholipase A1 and phospholipase B have a strong catalytic conversion effect on organic phosphorus, and the phosphorus-containing intermediates they generate are easier to convert into absorbable phosphorus, or are more easily absorbed by plants, which is conducive to the removal of organic phosphorus in water.

[0101] Table 1

[0102]

[0103]

[0104] (2) Effects of different types of phospholipases on total phosphorus removal

[0105] The concentration changes of total phosphorus in the simulated artificial wetland system are as follows Figure 4 As shown in Figure 2, the cumulative removal rate of total phosphorus changes as Figure 5 As shown. It can be seen that the removal rate of total phosphorus was the highest in the experimental group with the addition of phospholipase A1 and phospholipase B, and the removal rates of the groups with the addition of phospholipase A1 and phospholipase B were much higher than those of the experimental group without enzymes. This shows that both phospholipase A1 and phospholipase B can effectively promote the absorption and removal of total phosphorus by plants in simulated artificial wetlands, and the combined use effect is better.

[0106] Example 3

[0107] This example provides a study on the effect of different concentrations of phospholipase B on phosphorus removal in water.

[0108] In this embodiment, the plants used to simulate the artificial wetland system include water hyacinth, calamus and calamus. Figure 6 As shown, water hyacinth (3 to 5 pieces, height 6 to 7 cm), Acorus calamus (3 plants, height about 35 cm), and Acorus calamus (3 plants, height about 40 cm).

[0109] In this embodiment, the experiment was divided into three groups (including the control group), and 2L of natural lake water (water quality better than Class III water) and 3mL of plant nutrient solution (Melok plant nutrient solution universal type) were added to each group of simulated artificial wetland systems, and 10mL of 25mg / mL soybean lecithin emulsion was added to simulate wastewater containing organic phosphorus; the amount of phospholipase added in the three groups of experiments is as follows:

[0110] (1) Low-concentration enzyme experimental group: 24.4 g of the artificial wetland filler containing phospholipase B prepared in Example 1 (i.e., about 20 mg of phospholipase B was added);

[0111] (2) High-concentration enzyme experimental group: 73.2 g of the artificial wetland filler containing phospholipase B prepared in Example 1 (i.e., about 60 mg of phospholipase B was added);

[0112] (3) No enzyme experimental group: no phospholipase was added;

[0113] The experiment will start on May 3, 2023 and last for 2 weeks.

[0114] Result analysis:

[0115] (1) Removal effect of different concentrations of phospholipase B on organic phosphorus

[0116] The concentration changes of organic phosphorus in the simulated artificial wetland system are as follows Figure 7 The organic phosphorus concentration and removal rate are shown in Table 2.

[0117] like Figure 7As shown, the concentration of organic phosphorus in the water samples of the two simulated artificial wetland systems in which artificial wetland fillers containing phospholipase B were added first dropped rapidly, then stabilized and showed a fluctuating state. The concentration of organic phosphorus in the experimental group without enzyme addition (blank sample) first dropped steadily, then stabilized and showed a fluctuating state. By comparing these three, the following conclusion can be drawn: the artificial wetland filler of the present invention has a strong catalytic conversion effect on organic phosphorus.

[0118] Comparing the three groups of experiments, within one day of adding the artificial wetland filler, the concentration of organic phosphorus in the water sample of the high-concentration enzyme test group decreased by 89%, the concentration of organic phosphorus in the water sample of the low-concentration enzyme test group with a relatively small amount of added enzyme decreased by 64%, and the concentration of organic phosphorus in the blank sample (experimental group without enzyme addition) decreased by 15%. This shows that the rate of conversion of lecithin to soluble orthophosphate is proportional to the amount of filler added in the present invention within a certain range.

[0119] The concentration of organic phosphorus in the blank sample began to approach that of the first two experimental samples on the fifth day. This is because the plants themselves have various microorganisms that can participate in the decomposition of organic matter. Taking calamus as an example, there are rhizosphere bacteria, rhizosphere fungi, degrading bacteria, nitrifying bacteria and other microorganisms in the roots of calamus. These microorganisms can decompose organic matter and convert it into nutrients that can be used by plants. Organic phosphorus will also be converted in the process and slowly absorbed.

[0120] Starting from the sixth day of the experiment, the organic phosphorus concentrations in the three water bodies tended to a low and stable state with slight fluctuations. It is inferred that this is because the metabolism of plants and microorganisms caused some cells to enter the water bodies, and the phosphorus in these cells would also affect the changes in the organic phosphorus concentrations in the water bodies.

[0121] In the experimental group to which the artificial wetland filler of the present invention was added, the organic phosphorus entered a low level of organic phosphorus after 48 hours, while the blank control group without addition entered a relatively stable low level of organic phosphorus after one week. This also shows that adding the artificial wetland filler of the present invention can significantly improve the conversion and removal rate of organic phosphorus in this type of plant water purification system.

[0122] Table 2

[0123] Comparison of organic phosphorus removal rate Low concentration enzyme experimental group High concentration enzyme experimental group No enzyme experimental group Initial organic phosphorus (in terms of phosphorus P) (mg / L) 3.72 4.18 3.98 24h organic phosphorus (in terms of phosphorus P) (mg / L) 1.04 0.46 3.37 48h organic phosphorus (calculated as phosphorus P) (mg / L) 0.36 0.09 2.2 Organic phosphorus in one week (calculated as phosphorus P) (mg / L) 0.23 0.1 0.54 24-hour removal rate 72.04％ 89.00％ 15.33％ 48 hours removal rate 90.32％ 97.85％ 44.72％ Removal rate per week 93.82％ 97.61％ 86.43％ Average removal rate in the second week 95.65％ 97.94％ 95.98％

[0124] (2) Effect of different concentrations of phospholipase on total phosphorus removal

[0125] The concentration changes of total phosphorus in the simulated artificial wetland system are as follows Figure 8 As shown in Figure 2, the daily removal rate of total phosphorus changes as Fig. 9 As shown in Figure 2, the cumulative removal rate of total phosphorus changes as Fig.10 shown.

[0126] from Figure 8It can be seen that in the artificial wetland with high concentration of phospholipase, the removal rate of total phosphorus by plants is the highest. And in the same period of time, the removal rate is much higher than that of the artificial wetland without enzyme, which shows that phospholipase B will accelerate the absorption of phosphorus by plants in artificial wetlands.

[0127] from Fig. 9 It can be seen that the overall daily removal rate of the experimental sample with enzyme added is higher than that of the blank sample without enzyme added, which shows that phospholipase promotes the absorption of phosphorus by plants. The daily removal rate of total phosphorus is stable in a lower range, which shows that the removal of total phosphorus by artificial wetlands is a long process. This process is mainly through the absorption of phosphorus by plants. Therefore, from the perspective of daily removal rate, there is no significant difference between adding enzymes and not adding enzymes.

[0128] from Fig.10 It can be seen that the total phosphorus cumulative removal rate in 2 weeks, that is, the total removal rate, is significantly higher than that of the group without enzyme addition, whether it is high-concentration or low-concentration phospholipase. This shows that the addition of phospholipase is not only beneficial to the catalytic conversion of organic phosphorus, but also can promote the absorption and removal of total phosphorus to a certain extent.

[0129] (3) Effects of different phospholipase concentrations on plant height changes in simulated artificial wetland systems

[0130] Simulating plant height changes in artificial wetland systems Figures 11 to 13 As shown in the figure, the height of the three plants increased, but the height increment of the experimental group with phospholipase added was significantly higher than that of the blank sample without phospholipase. The height of water hyacinth and Acorus calamus increased more significantly in the high concentration phospholipase group. This shows that the addition of phospholipase is conducive to the catalytic conversion of organic phosphorus, which is easier to be absorbed by plants and the plants grow better. Among them, the promotion effect on Acorus calamus is better, with the increments of low and high concentration groups reaching 16.02cm and 16.47cm respectively, which is nearly twice the blank sample of 8.43cm.

Claims

1. Application of phospholipase in promoting the removal of phosphorus from water bodies, the application comprising adding phospholipase to a wetland or artificial wetland system, wherein the phospholipase comprises any one or more of phospholipase A1, phospholipase A2, phospholipase B, phospholipase C and phospholipase D.

2. The use of the phospholipase according to claim 1 in promoting the removal of phosphorus from water, characterized in that: The phospholipase includes phospholipase A1 and / or phospholipase B.

3. Use of the phospholipase according to claim 1 or 2 in promoting the removal of phosphorus from water, characterized in that: The application includes: using nanospheres as filler matrix, loading phospholipase, and using them as artificial wetland fillers after being wrapped with sodium alginate.

4. The use of the phospholipase according to claim 3 in promoting the removal of phosphorus from water bodies, characterized in that: The artificial wetland filler also includes CaCl2.

5. The use of the phospholipase according to claim 4 in promoting the removal of phosphorus from water bodies, characterized in that: The artificial wetland filler also includes indoleacetic acid, vitamin B1, vitamin B6, calcium, iron, zinc and magnesium.

6. The method for preparing the artificial wetland filler according to claim 4, characterized in that: The method comprises: adding nanospheres into a phospholipase solution, shaking and then standing to obtain catalytic nanospheres; adding the catalytic nanospheres into a sodium alginate solution, stirring until the surface of the catalytic nanospheres is covered with sodium alginate; and then putting the catalytic nanospheres into a calcium chloride solution, embedding and then taking them out.

7. The method for preparing the artificial wetland filler according to claim 5, characterized in that: The method comprises: adding nanospheres to a phospholipase solution, shaking and then standing to obtain catalytic nanospheres; adding the nanospheres to a mixed auxiliary liquid containing indoleacetic acid, vitamin B1, vitamin B6, calcium, iron, zinc and magnesium, shaking and then standing to obtain auxiliary nanospheres; adding the catalytic nanospheres and auxiliary nanospheres to a sodium alginate solution, stirring until the surfaces thereof are covered with sodium alginate; and then putting them into a calcium chloride solution, embedding and then taking them out.

8. An artificial wetland filler for promoting phosphorus removal from water, characterized in that: The artificial wetland filler comprises: Nanospheres, Phospholipase, wherein the phospholipase includes any one or more of phospholipase A1, phospholipase A2, phospholipase B, phospholipase C and phospholipase D, Sodium alginate, Calcium chloride, The phospholipase is loaded on the nanospheres, and the sodium alginate wraps the nanospheres loaded with the phospholipase.

9. The artificial wetland filler for promoting phosphorus removal in water according to claim 8, characterized in that: The artificial wetland filler also includes: indoleacetic acid, vitamin B1, vitamin B6, calcium, iron, zinc and magnesium.

10. An artificial wetland system, characterized in that: The filler in the artificial wetland system includes the artificial wetland filler for promoting the removal of phosphorus from water bodies as described in claim 8 or 9.

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