An intelligent pH-sensing algistatic flavone sustained-release microsphere and a preparation method thereof

Abstract

The application discloses an acid-base intelligent sensing algae-inhibiting flavone slow-release microsphere and a preparation method thereof. The effective algae-inhibiting substance in the acid-base intelligent sensing algae-inhibiting flavone slow-release microsphere is 5,4'-dihydroxy flavone, and the acid-base sensing material is stearic acid. The flavone-stearic acid-sodium alginate mixed solution is dropped into the chitosan-calcium chloride mixed solution for fixation, then the microspheres are filtered out and freeze-dried, so that the acid-base intelligent sensing algae-inhibiting flavone slow-release microsphere is obtained. The material used in the application has good aquatic ecological safety, and the prepared acid-base intelligent sensing algae-inhibiting flavone slow-release microsphere can dynamically control the release rate of 5,4'-dihydroxy flavone in response to the change of the water body acid-base value, accurately inhibits the growth of water bloom algae, especially the growth of Microcystis aeruginosa, and can effectively control the outbreak of algae bloom. Compared with the prior art, the effective utilization rate of the algae-inhibiting agent is improved, the algae-inhibiting duration is more persistent, and the application is particularly suitable for the prevention and control of cyanobacterial bloom.

Description

Technical Field

[0001] This invention belongs to the field of water pollution control, and in particular relates to a pH-sensitive, algae-inhibiting, slow-release microsphere and its preparation method. Background Technology

[0002] Against the backdrop of eutrophication and global warming, harmful cyanobacteria are rapidly proliferating and forming algal blooms, which not only disrupt the balance of ecosystems but also endanger the sustainable development of inland and nearshore sea areas, causing huge losses to aquaculture and even threatening human health. In recent years, due to the enormous harm caused by cyanobacterial blooms, precise, efficient, long-term dynamic, and low-ecological-risk algal control technologies have received widespread attention from researchers. The applicant's patent application in 2011 (patent number: ZL2012102108685, titled: Preparation method and application of the barley allelochemical algal inhibitory isomer, cycladine) clearly points out that the flavonoid lignin cycladine in barley straw plays a major role in algal inhibition. At the same time, numerous studies have shown that flavonoids are allelochemicals with good algal inhibitory effects, among which 5,4'-dihydroxyflavone has a stronger algal inhibitory effect and has the potential to be used as an algal inhibitor for algal bloom control.

[0003] In practical applications of algae suppression, directly introducing flavonoids as algaecides into water bodies can easily lead to excessively high local flavonoid concentrations, interfering with the normal growth of non-target organisms and adversely affecting the aquatic ecosystem. Furthermore, the algae-suppressing effect is short-lived. In addition, algal blooms in real-world water bodies are influenced by numerous environmental factors, with pH being a crucial one. Numerous studies have shown that cyanobacteria exhibit higher growth rates in slightly alkaline water (pH 8-9), are more prone to rapid proliferation and algal blooms, and can even regulate the pH of the water to a suitable level for their growth. Moreover, when algal density is high, large amounts of carbon dioxide are absorbed for photosynthesis, further increasing the water's pH. Therefore, the pH of the water body is closely related to the growth of cyanobacteria. Currently available slow-release algaecides / microspheres can only achieve slow release of algaecides in water at a certain rate. They cannot dynamically regulate the release rate of algaecides in response to changes in the growth rate of cyanobacteria caused by changes in the pH of the water. This results in insufficient utilization of algaecides, reduced algaecide duration, and an increased probability of side effects on the water.

[0004] To address the aforementioned shortcomings, this invention, based on previous research and drawing on intelligent controlled-release systems for pesticides, develops pH-sensitive, algae-inhibiting flavonoid-releasing microspheres. These microspheres can sense and respond to changes in water pH to dynamically and precisely regulate the release rate of algae-inhibiting flavonoids, thereby improving the utilization rate of algae inhibitors, extending the duration of algae inhibition, and enhancing the ecological safety of algae inhibitors. This contributes to achieving intelligent, precise, and dynamic algae control. Summary of the Invention

[0005] The purpose of this invention is to address the shortcomings of existing technologies by providing a pH-sensitive, algae-inhibiting, sustained-release microsphere and its preparation method.

[0006] The present invention is achieved through the following technical solution: a pH-sensitive algae-inhibiting flavonoid slow-release microsphere, wherein the effective algae-inhibiting substance is 5,4'-dihydroxyflavone, the pH-sensing material is stearic acid, and the encapsulation material includes sodium alginate, chitosan and calcium chloride, with an encapsulation rate of 60-90%.

[0007] The preparation method of the above-mentioned pH-sensitive algae-inhibiting flavonoid sustained-release microspheres includes the following steps:

[0008] (1) Stearic acid and sodium alginate were dissolved in anhydrous ethanol and deionized water, respectively, to obtain an anhydrous ethanol solution of stearic acid and an aqueous solution of sodium alginate. 5,4'-dihydroxyflavone powder, anhydrous ethanol solution of stearic acid and aqueous solution of sodium alginate were mixed to prepare a flavonoid-stearic acid-sodium alginate mixed solution, wherein the concentrations of 5,4'-dihydroxyflavone, stearic acid and sodium alginate were 15-20 g / L, 5-10 g / L and 15-25 g / L, respectively, and the volume fraction of anhydrous ethanol was 15-20%.

[0009] (2) Chitosan and anhydrous calcium chloride powder are dissolved in a 1% acetic acid solution to prepare a chitosan-calcium chloride mixed solution, wherein the concentrations of chitosan and calcium chloride are 20-30 g / L and 10-20 g / L, respectively.

[0010] (3) The flavonoid-stearic acid-sodium alginate mixed solution prepared in step (1) is added dropwise to the chitosan-calcium chloride mixed solution prepared in step (2) for fixation. Stirring is done while adding the solution to obtain shaped microspheres, which are then collected by filtration using a sieve.

[0011] (4) The microspheres obtained in step (3) are repeatedly rinsed with deionized water and then freeze-dried to obtain pH-sensitive algae-inhibiting flavonoid sustained-release microspheres.

[0012] Further, in step (1), the 5,4'-dihydroxyflavone powder, the anhydrous ethanol solution of stearic acid and the aqueous solution of sodium alginate are thoroughly mixed using an electric stirrer.

[0013] Further, in step (3), the flavonoid-stearic acid-sodium alginate mixed solution is added dropwise to the chitosan-calcium chloride mixed solution for fixation using a syringe, while stirring with a magnetic stirrer at 500 rpm for 30 minutes. The formed microspheres are then collected by filtration through a sieve.

[0014] Further, in step (4), the microspheres are repeatedly rinsed with deionized water and then placed in a freeze dryer to dry for 12 hours to obtain pH-sensitive algae-inhibiting flavonoid sustained-release microspheres.

[0015] Further, the following preferred parameters may be used, but are not limited to: in the flavonoid-stearic acid-sodium alginate mixed solution, the concentrations of 5,4'-dihydroxyflavone, stearic acid and sodium alginate are 20 g / L, 5 g / L and 15 g / L, respectively, and the volume fraction of anhydrous ethanol is 17.5%; in the chitosan-calcium chloride mixed solution, the concentrations of chitosan and calcium chloride are 20 g / L and 10 g / L, respectively.

[0016] The prepared pH-sensitive, slow-release microspheres of algal-inhibiting flavonoids can dynamically regulate the release rate of the effective algal-inhibiting substance 5,4'-dihydroxyflavone in response to changes in water pH. This can be applied to the dynamic and precise control or treatment of algal blooms in lakes.

[0017] The present invention has the following beneficial effects:

[0018] 1. This invention encapsulates the algae inhibitor 5,4'-dihydroxyflavone and the pH-sensing material stearic acid within a sodium alginate-chitosan backbone. This allows the algae inhibitor to sense and respond to changes in pH in the environment, precisely and dynamically controlling the release rate of the algae inhibitor 5,4'-dihydroxyflavone, thereby improving the effective utilization rate of the algae inhibitor and avoiding waste; extending the algae-inhibiting effect and saving manpower; and improving the ecological safety of the algae inhibitor and reducing toxic side effects.

[0019] 2. The algaecide 5,4'-dihydroxyflavone used in the preparation of the pH-sensitive, algaecide-inhibiting, sustained-release microspheres of this invention is an allelochemical, easily degradable, with no residue risk, and will not cause secondary pollution; the pH-sensing material, stearic acid, has no obvious toxic side effects on aquatic organisms; the encapsulation materials, chitosan and sodium alginate, are natural polysaccharides with high ecological safety. These materials are convenient to use, widely available, and inexpensive, making them suitable for large-scale promotion. Attached Figure Description

[0020] The accompanying drawings, which form part of this invention, are used to provide a further understanding of the invention. The illustrative embodiments of the invention and their descriptions are used to explain the invention and do not constitute an improper limitation of the invention.

[0021] Figure 1 The release rate of 5,4'-dihydroxyflavone from the pH-sensitive algae-inhibiting flavonoid sustained-release microspheres in this embodiment of the invention is shown in different pH values ​​(7, 7.5, 8, 8.5, 9).

[0022] Figure 2(a), (b), and (c) represent the algae inhibition rates of the pH-sensitive algae-inhibiting flavonoid slow-release microspheres and the 5,4'-dihydroxyflavone solution in the embodiments of the present invention under pH conditions of 7, 8, and 9, respectively. Detailed Implementation

[0023] The present invention will be further illustrated below through examples.

[0024] Example 1

[0025] The preparation process parameters of the pH-sensitive algae-inhibiting flavonoid sustained-release microspheres were optimized by orthogonal experiments. Sodium alginate, chitosan, calcium chloride, 5,4'-dihydroxyflavone, stearic acid and anhydrous ethanol were used as the six elements of the orthogonal experiments, and three levels were set for each element, as shown in Table 1.

[0026] Table 1 Orthogonal Experimental Design

[0027]

[0028] Sodium alginate powder was weighed and dissolved in deionized water to prepare aqueous solutions with concentrations of 15 g / L, 20 g / L, and 25 g / L. Stearic acid was weighed and dissolved in anhydrous ethanol to prepare an anhydrous ethanol solution of stearic acid. Using an electric stirrer, 5,4'-dihydroxyflavone powder, the anhydrous ethanol solution of stearic acid, and the aqueous solution of sodium alginate were mixed to prepare flavonoid-stearic acid-sodium alginate mixed solutions of different concentrations, wherein the concentrations of 5,4'-dihydroxyflavone were 15 g / L, 17.5 g / L, and 20 g / L, the concentrations of stearic acid were 5 g / L, 7.5 g / L, and 10 g / L, and the volume fractions of anhydrous ethanol were 15%, 17.5%, and 20%. Chitosan and anhydrous calcium chloride were weighed and dissolved in a 1% (v / v) deionized water solution of acetic acid to prepare chitosan-calcium chloride mixed solutions of different concentrations. The concentrations of chitosan in the mixed solutions were 20 g / L, 25 g / L, and 30 g / L, and the concentrations of calcium chloride were 10 g / L, 15 g / L, and 20 g / L, respectively. The prepared flavonoid-stearic acid-sodium alginate mixed solution was added dropwise to the chitosan-calcium chloride mixed solution using a syringe for fixation. The mixture was stirred at 500 rpm using a magnetic stirrer for 30 minutes. After stirring, the formed microspheres were collected by filtration through a sieve, rinsed repeatedly with deionized water, and then dried in a freeze dryer for 12 hours to obtain pH-sensitive algae-inhibiting flavonoid slow-release microspheres. The optimal preparation scheme for the pH-sensitive, algae-inhibiting, flavonoid-releasing microspheres was as follows: 15 g / L sodium alginate, 10 g / L calcium chloride, 20 g / L chitosan, 20 g / L 5,4'-dihydroxyflavone solution, 5 g / L stearic acid, and 17.5% (v / v) anhydrous ethanol. Under these optimal conditions, the encapsulation efficiency of the pH-sensitive, algae-inhibiting, flavonoid-releasing microspheres was 89.05%. Conventional algae inhibition experiments showed that the obtained product exhibited significant algae-inhibiting activity.

[0029] Example 2

[0030] BG11 medium suitable for cyanobacterial growth was prepared. The pH of the BG11 medium was adjusted to 7.0, 7.5, 8.0, 8.5, and 9.0 using 0.1 mol / L hydrochloric acid and sodium hydroxide. Then, the pH-sensitive algae-inhibiting flavonoid slow-release microspheres prepared under the optimal conditions in Example 1 were immersed in 8 mL of BG11 medium at different pH values. The microspheres were placed in a constant temperature incubator at 25°C to detect the release rate of 5,4'-dihydroxyflavone at different pH values. The concentration of 5,4'-dihydroxyflavone in the medium was detected daily for the first 10 days. From day 11 to 40, the concentration was detected every 5 days. After each concentration test, the same volume of BG11 medium with the same pH value was added to maintain a consistent release environment.

[0031] Experimental results are as follows Figure 1 As shown, the data indicates that the release rate of 5,4'-dihydroxyflavone from the pH-sensitive algae-inhibiting flavonoid microspheres is significantly lower when the pH is below 8 than when the pH is between 8 and 9. This demonstrates that the microspheres can dynamically and precisely regulate the release rate of the algae-inhibiting agent 5,4'-dihydroxyflavone by sensing changes in the pH of the aquatic environment, thereby improving the effective utilization rate of 5,4'-dihydroxyflavone.

[0032] Example 3

[0033] Microcystis aeruginosa (FACHB 905) was purchased from the Institute of Hydrobiology, Chinese Academy of Sciences, and cultured in BG11 medium under a constant temperature and light incubator at 25±1℃, 3000 lux light intensity, and a light-dark ratio of 12:12. 50 mL of BG11 medium, 0.1 mL of Microcystis aeruginosa in its logarithmic growth phase, and an appropriate amount of pH-sensitive algicidal flavonoid slow-release microspheres from Example 1 were added to a 100 mL Erlenmeyer flask to achieve an initial algal density of 5 × 10⁻⁶. 5 The final concentration of the pH-sensitive, slow-release algae-inhibiting flavonoid microspheres was 0.15 g / L. The concentration of 5,4'-dihydroxyflavone in the treatment group (directly adding 5,4'-dihydroxyflavone solution) was 1 mg / L. A blank control group (no algae-inhibiting agent added) was used. The BG11 culture medium used in the pH-sensitive, slow-release algae-inhibiting flavonoid microsphere treatment group, the 5,4'-dihydroxyflavone solution treatment group, and the blank control group (no algae-inhibiting agent added) was adjusted to pH 7, 8, and 9 respectively using 0.1 mol / L hydrochloric acid and sodium hydroxide. The culture media were then placed in a 25°C incubator to monitor changes in algae inhibition rate. The pH of BG11 was measured daily during the experiment, and if changes occurred, it was readjusted using hydrochloric acid and sodium hydroxide. The experiment lasted 20 days. Algal cells were sampled and tested every 2 days for the first 10 days and every 5 days for the following 10 days.

[0034] Formula for calculating algae inhibition rate: Algae inhibition rate % = (C c -C s )×100 / C c ;

[0035] Where C s To treat algal density, C c The algal density represents the blank control group.

[0036] Experimental results are as follows Figure 2 As shown, the data indicates that compared with low pH (7-8), the algae inhibition rate of directly adding 5,4'-dihydroxyflavone solution decreased significantly under pH 9 conditions, failing to maintain long-term high-efficiency algae inhibition. However, pH-sensitive algae-inhibiting flavonoid slow-release microspheres can maintain long-term high-efficiency algae inhibition under different pH conditions (7-9). This is because the pH-sensitive algae-inhibiting flavonoid slow-release microspheres have pH-sensing response release characteristics. When the pH increases, the growth rate of Microcystis aeruginosa increases, and the pH-sensitive algae-inhibiting flavonoid slow-release microspheres can release 5,4'-dihydroxyflavone at an accelerated rate, maintaining a high-efficiency algae inhibition effect under higher pH conditions, and has the potential for precise and dynamic algae control.

[0037] Although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art can still modify the technical solutions described in the foregoing embodiments or make equivalent substitutions for some of the technical features. Any modifications, substitutions, or improvements made within the spirit and principles of the present invention should be included within the scope of protection of the present invention. Therefore, the content of this specification should not be construed as a limitation of the present invention.

Claims

1. A method for preparing pH-sensitive, algae-inhibiting, sustained-release microspheres, characterized in that, Includes the following steps: (1) Stearic acid and sodium alginate were dissolved in anhydrous ethanol and deionized water, respectively, to obtain an anhydrous ethanol solution of stearic acid and an aqueous solution of sodium alginate. 5,4'-dihydroxyflavone powder, anhydrous ethanol solution of stearic acid and aqueous solution of sodium alginate were mixed to prepare a flavonoid-stearic acid-sodium alginate mixed solution, wherein the concentrations of 5,4'-dihydroxyflavone, stearic acid and sodium alginate were 15-20 g / L, 5-10 g / L and 15-25 g / L, respectively, and the volume fraction of anhydrous ethanol was 15-20%; the 5,4'-dihydroxyflavone was used as an effective algae-inhibiting substance; the stearic acid was used as an acid-base sensing material; and the sodium alginate was used as an embedding material. (2) Chitosan and anhydrous calcium chloride powder are dissolved in a 1% acetic acid solution to prepare a chitosan-calcium chloride mixed solution, wherein the concentrations of chitosan and calcium chloride are 20-30 g / L and 10-20 g / L, respectively; the chitosan and calcium chloride are used as embedding materials. (3) The flavonoid-stearic acid-sodium alginate mixed solution prepared in step (1) is added dropwise to the chitosan-calcium chloride mixed solution prepared in step (2) for fixation. Stirring is done while adding the solution to obtain shaped microspheres, which are then collected by filtration using a sieve. (4) The microspheres obtained in step (3) are repeatedly rinsed with deionized water and then freeze-dried to obtain pH-sensitive algae-inhibiting flavonoid slow-release microspheres, which can dynamically regulate the release rate of the effective algae-inhibiting substance 5,4'-dihydroxyflavone in response to changes in the pH of the water.

2. The preparation method according to claim 1, characterized in that, In step (1), the 5,4'-dihydroxyflavone powder, anhydrous ethanol solution of stearic acid and aqueous solution of sodium alginate are thoroughly mixed using an electric stirrer.

3. The preparation method according to claim 1, characterized in that, In step (3), the flavonoid-stearic acid-sodium alginate mixed solution is added dropwise to the chitosan-calcium chloride mixed solution for fixation using a syringe. At the same time, a magnetic stirrer is used to stir the solution at a speed of 500 rpm for 30 minutes. After stirring, the formed microspheres are collected by filtration through a sieve.

4. The preparation method according to claim 1, characterized in that, In step (4), the microspheres are repeatedly rinsed with deionized water and then placed in a freeze dryer to dry for 12 hours to obtain pH-sensitive algae-inhibiting flavonoid sustained-release microspheres.

5. The preparation method according to claim 1, characterized in that, In the flavonoid-stearic acid-sodium alginate mixed solution, the concentrations of 5,4'-dihydroxyflavone, stearic acid, and sodium alginate were 20 g / L, 5 g / L, and 15 g / L, respectively, and the volume fraction of anhydrous ethanol was 17.5%; in the chitosan-calcium chloride mixed solution, the concentrations of chitosan and calcium chloride were 20 g / L and 10 g / L, respectively.

6. A pH-sensitive, algae-inhibiting, sustained-release microsphere, characterized in that, It is prepared by the preparation method described in any one of claims 1-5.

7. The application of the pH-sensitive algae-inhibiting flavonoid slow-release microspheres as described in claim 6 in the dynamic and precise control or treatment of algal blooms in lakes.

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