Preparation of amylopectin and gelatin moldable sponge and related application of amylopectin and gelatin moldable sponge in micro-plastic removal

Abstract

The invention discloses preparation of amylopectin and gelatin plastic sponge and related application of the amylopectin and gelatin plastic sponge in micro-plastic removal, and belongs to the field of micro-plastic removal. According to the technical scheme adopted by the invention, the preparation method comprises the following steps: preparing starch and gelatin semi-interpenetrating cross-linked sponge, and respectively carrying out cross-linking reaction on starch with the concentration of 4% and gelatin with the concentration of 3%. The preparation method comprises the following steps: uniformly stirring at the initial condition that the ratio of the two is 1: 1 after crosslinking, quickly freezing at-80 DEG C for 48 hours, then freeze-drying, and then extruding and adsorbing sponges prepared by optimizing various conditions with micro-plastic solutions in different food environments respectively. The efficient micro-plastic removal method (the enrichment efficiency is about 90% or above) is established, the method is expanded to be applied to samples containing a large number of small particles and organic matter such as seawater, soil and take-out vegetable soup, and meanwhile the problem that micro-plastic in a complex matrix is difficult to enrich is solved.

Description

Technical Field

[0001] The present invention relates to the removal of micro / nano plastics in an environmental system, and in particular to a method for efficiently removing micro plastics from an environmental matrix using a semi-interpenetrating cross-linked sponge based on starch and gelatin, belonging to the technical field of micro plastic removal and application. Background Art

[0002] Every year, nearly 320 million tons of plastic waste are gradually degraded into smaller plastic particles (<5 mm) due to weathering and seawater erosion, namely micro / nano plastics. As emerging pollutants, they are ubiquitous in the environment (sea ports, rivers and soil), and their harmful effects on ecosystems and human health have become a matter of great concern to the world. For example, polystyrene (PS) and polymethyl methacrylate (PMMA) are widely used in food packaging and dental biomedical materials (dental clinics, dental laboratories and artificial teeth). Currently, they are found in wheat root crowns, Antarctic krill, and even in human organs and tissues. Due to their strong hydrophobicity and adsorption activity, micro / nano plastics often form "protein crowns" with protein molecules, become carriers of various pollutants, and eventually enter the food chain, causing more extensive toxicity to organisms and humans. Therefore, it is urgent to provide a method to remove micro / nano plastic particles from the environment and even our food.

[0003] In the field of methods for removing micro / nano plastics, methods such as magnetic particle adsorption, chemical flocculant precipitation and nanorobot sweeping are currently widely reported. However, the above methods are subject to many limitations in practical applications. Magnetic particle adsorption is limited to the removal of nanoplastics in the environment, which limits the application range of its size. Although the nanorobot sweeping method is greener than the chemical flocculant precipitation method, its low flux, high cost and harsh application conditions still limit its industrial application. Furthermore, composite sponges have shown initial results in effectively removing micro / nano plastics. Chitin-rich sponges cross-linked with graphene oxide can remove micro- and nano-plastics from water, respectively. However, this sponge is potentially toxic due to the doping of graphene oxide. For greener and non-toxic applications, similar sponges were reported to be prepared using plant protein as raw materials, but the high cost still limits their application. Therefore, research is committed to developing low-cost, simple and sustainable tools or methods to remove micro / nano plastics from the environment and food matrices.

[0004] Starch is a cheap natural polymer widely found in plant seeds, tubers and roots. Due to its degradability and water absorption capacity, starch is widely used as an ideal green chemical material for the manufacture of hemostatic and antibacterial sponges. However, the poor mechanical strength of starch has hindered the further development of starch materials. Gelatin is a protein composed of multiple amino acids connected by amide bonds, which has fracture resistance and good elasticity. It is considered to be a good support for improving the mechanical strength of various materials. Although both ingredients show the advantage of degradability, sponges prepared from starch and gelatin are insufficiently studied, resulting in a lack of corresponding technology for the removal of micro / nano plastics. At the same time, there are no public reports on the preparation of plastic starch-gelatin cross-linked sponges and their application in the removal of micro / nano plastics in environmentally contaminated samples. Summary of the invention

[0005] In view of the problems existing in the prior art, the present invention provides a preparation method of a plastic sponge of amylopectin and gelatin and its application in the removal of microplastics.

[0006] In order to achieve the above object, the present invention adopts the following technical solutions: A preparation method of a plastic sponge of amylopectin and gelatin and its application in microplastic removal, comprising the following steps: S1: Preparation of semi-interpenetrating starch and gelatin sponge: starch concentration of 4% and gelatin concentration of 3% were cross-linked, mixed evenly at an initial ratio of 1:1, and freeze-dried at -80℃ for 48h; S2: Adsorption treatment of starch-gelatin sponge and microplastics was performed. The sponge prepared after optimizing various conditions was squeezed and adsorbed with microplastic solutions in different food environments; S3: To investigate the state of the adsorption mechanism of microplastics and sponges, 50 mg of sponges were used to adsorb 5 mL of 25 mg / mL microplastic solutions at different pH and sodium chloride concentrations; S4: To evaluate the degradability of starch-gelatin sponges, different masses of sponges were cultured with corresponding bacterial colonies in different types of culture media of the same mass. The OD of the bacterial colonies at different time points was measured. 600 Determine the biocompatibility of the sponge.

[0007] Preferably, in step S1, the concentration of the starch solution is fixed at 4%, and the concentration of the starch solution is optimized according to different preparation conditions.

[0008] Preferably, the starch suspension in step S1 is prepared by gelatinizing waxy corn starch at 92° C. (500 rpm) for 1 h.

[0009] Preferably, in step S1, the gelatin solution is continuously stirred at 70° C. and formaldehyde (0.35%, v / v) is slowly added to the above system within 1 minute for chemical cross-linking.

[0010] Preferably, the operation method in step S2 is divided into a three-way valve extrusion method and a chromatography filtration method.

[0011] Preferably, the adsorption mechanism of the sponge and sodium chloride in step S3 is explored by repeatedly squeezing and adsorbing 50 mg of sponge with 40 mL of a 25 mg / L fluorescent microplastic solution (PS: 100 nm; PMMA: 5 μm) through a kinetic experiment.

[0012] Preferably, in step S4, regarding the enzyme degradation determination step, a certain amount of sponge is taken, and the sponge after removing 5 μm and 100 nm is reacted in a system containing α-amylase (3,000 U) and saccharifying enzyme. Finally, the concentration of glucose in the system at different time points is determined.

[0013] Preferably, in step S4, regarding the biocompatibility evaluation, the mass fraction of the sponge is (0.05%, 1%, 2% and 6%, w / v), the reaction system is 1 mL of Pichia. Pastoris in the logarithmic growth phase per 30 mL of YPD medium, 1 mL of Escherichia coli in the logarithmic growth phase per 30 mL of LB medium, and 1 mL of Lactic acid bacteria in the logarithmic growth phase per 30 mL of MRS medium, and the reaction temperature is 37°C.

[0014] Preferably, the amount of sponge used in the glucose determination in step S4 is 200 mg, the degradation system is 20 mL of physiological saline containing α-amylase (3,000 U) and saccharifying enzyme (5,000 U), and the reaction temperature is 37°C.

[0015] Compared with the prior art, the present invention provides a preparation method for a plastic sponge of amylopectin and gelatin and its application in the removal of microplastics, which has the following beneficial effects: the present invention establishes an efficient method for the removal of microplastics (enrichment efficiency is about 90% or more), and expands its application to samples such as seawater, soil, and takeaway vegetable soup containing a large number of small particles and organic matter. In this process, the problem of the difficulty of enriching microplastics in complex matrices is solved, the operating cost is reduced, the method of replacing high-cost filtering devices such as filter membranes with sponges is expanded, and the problem of density flotation and chemical precipitation methods being unsuitable for small particles and secondary pollution is avoided. The present invention not only provides strong technical support for the enrichment and adsorption of plastic pollution, but also provides a feasible solution for new products of low-cost filtering phases in environmental, food and other departments to protect the life, health and safety of consumers. BRIEF DESCRIPTION OF THE DRAWINGS

[0016] Figure 1 A diagram showing the steps for preparing starch-gelatin sponge in the present invention; Figure 2 is the Fourier transform infrared spectra of gelatin, starch and sponge in the present invention; Figure 3 Schematic diagram of various sponges prepared with different starch / gelatin ratios in the present invention; Figure 4 It is a columnar schematic diagram of the expansion rate of the sponge at different gelatin concentrations in the present invention; Figure 5 Schematic diagram of the porosity of the sponge at different gelatin concentrations (4% and 5%) in the present invention; Figure 6 This is a schematic diagram of the sponge preparation of the present invention; Figure 7 This is a schematic diagram of placing starch-gelatin sponge on a flower in the present invention; Figure 8 Schematic diagram of the steps of removing micro / nano plastics from water by starch-gelatin sponge in the present invention; Fig. 9 Schematic diagram of the effect of different concentrations and types of micro / nano plastics on removal efficiency in the present invention; Fig.10 Schematic diagram of the effect of NaCl concentration and pH value on the removal ability of micro / nano plastics in the present invention; Fig.11 Representative SEM images of microplastics adsorbed by sponges prepared at different ratios in the present invention; Fig.12 This is a schematic diagram of the mechanism of starch-gelatin sponge adsorbing microplastics in the present invention; Fig.13 This is a diagram showing the application of starch-gelatin sponge in the present invention in removing microplastics from actual samples; Fig.14 It is a schematic diagram of the growth curves of lactic acid bacteria, Escherichia coli and Pichia pastoris in the present invention after being cultured with different amounts of sponges and sponges after microplastics have been removed for different periods of time; Fig.15 Schematic diagram of glucose degradation of the sponge, the sponge with 5 μm microplastics removed, and the sponge with 100 nm plastics removed in the present invention; Fig.16 Schematic diagram of starch degradation under α-amylase treatment in the present invention; Fig.17 Schematic diagram of the natural degradation of starch-gelatin sponge in soil in the present invention. DETAILED DESCRIPTION

[0017] The technical solutions in the embodiments of the present invention will be described clearly and completely below in conjunction with the drawings in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, rather than all the embodiments.

[0018] In the description of the present invention, it is necessary to understand that the terms "upper", "lower", "front", "back", "left", "right", "top", "bottom", "inside", "outside", etc., indicating the orientation or position relationship are based on the orientation or position relationship shown in the drawings, and are only for the convenience of describing the present invention and simplifying the description, rather than indicating or implying that the device or element referred to must have a specific orientation, be constructed and operated in a specific orientation, and therefore cannot be understood as a limitation on the present invention.

[0019] Embodiment 1: Reference Figure 1-17 , a preparation of a plastic sponge of amylopectin and gelatin and its application in microplastic removal, comprising the following steps: S1: Preparation of semi-interpenetrating starch and gelatin sponge: starch concentration of 4% and gelatin concentration of 3% were cross-linked, mixed evenly at an initial ratio of 1:1, and freeze-dried at -80℃ for 48h; S2: Adsorption treatment of starch-gelatin sponge and microplastics was performed. The sponge prepared after optimizing various conditions was squeezed and adsorbed with microplastic solutions in different food environments; S3: To investigate the state of the adsorption mechanism of microplastics and sponges, 50 mg of sponges were used to adsorb 5 mL of 25 mg / mL microplastic solutions at different pH and sodium chloride concentrations; S4: To evaluate the degradability of starch-gelatin sponges, different masses of sponges were cultured with corresponding bacterial colonies in different types of culture media of the same mass. The OD of the bacterial colonies at different time points was measured. 600 Determine the biocompatibility of the sponge.

[0020] Furthermore, in step S1, the concentration of starch solution was fixed at 4%, and the concentration of starch solution was optimized for different preparation conditions, including gelatin concentration (3%, 4%, 5%, 6% and 7%, v / v) and starch-gelatin ratio (1:1, 1:2, 1:3 and 1:4, v / v), such as Figure 3-5 As shown, the optimal ratio of starch and gelatin when preparing sponge is determined by porosity and water absorption, and is calculated as shown in formulas (1-1) and (1-2).

[0021] Porosity (%) = (1-1) Among them, m t and m 0represents the weight before and after ethanol absorption, ρ represents the density of ethanol, v 0 represents the volume of ethanol absorbed.

[0022] Water absorption (%) = (1-2) Among them, m t and m 0 Indicates the weight after and before water absorption.

[0023] Furthermore, the starch suspension in step S1 is prepared by gelatinizing waxy corn starch at 92°C (500rpm) for 1h. The pH value of the gelatinized starch solution is adjusted to 10 with 0.1mol / L NaOH solution, and then epichlorohydrin (500rpm) (1%, v / v) is added dropwise to the above solution within 5 minutes. The system solution is kept at 92°C for chemical crosslinking for 1h to complete the crosslinking process. The chemical crosslinking process is shown in the figure below. Figure 1 shown.

[0024] Furthermore, the gelatin solution in step S1 was chemically cross-linked by slowly adding formaldehyde (0.35%, v / v) to the above system under continuous stirring at 70°C for 1 minute. The solution was cross-linked for 30 minutes under continuous stirring (500 rpm) at 70°C. The chemical cross-linking mechanism is as follows Figure 1 c. The sponge with the best ratio in step (1) is characterized by its pore and ultra-light weight characteristics, such as Figure 2 , Figure 6 and Figure 7 shown.

[0025] Embodiment 2: Reference Figure 1-17 , which is basically the same as Example 1, and further, the operation method in step S2 is divided into a three-way valve extrusion method and a chromatography filtration method. The sponges prepared after optimizing various conditions are squeezed and adsorbed with microplastic solutions in different food environments.

[0026] The three-way valve extrusion method is as follows: first, the soil and takeaway vegetable soup are naturally settled to prepare a suspension and the supernatant is taken. Then, the micro-nano plastic solution is diluted to 25 mg / L with tap water, sea water, soil water and takeaway vegetable soup supernatant. Then, the starch-gelatin sponge is placed in a syringe connected to a three-way valve, and 5 mL of the prepared micro-nano plastic solution is sucked in respectively. The operation process is as follows: Figure 8 As shown, after the solution was repeatedly squeezed out in a three-way valve for removal, the removal rate was determined based on the fluorescence intensity.

[0027] The operation steps of the chromatography filtration method are as follows: fill the prepared starch-gelatin sponge into the chromatography column, prepare the suspension after natural sedimentation of soil and take-out vegetable soup and take the supernatant, and then use tap water, sea water, soil water and take-out vegetable soup supernatant to prepare a micro-nano plastic solution with a concentration of 25 mg / L, take 5 mL of the above respectively for filtration, collect the filtered liquid at the bottom, and measure the fluorescence value. The removal rates of the above two methods are calculated according to formula (2-1).

[0028] Q = (C 0 —C e ) / C 0 (2-1) Among them, C e and C 0 They represent the fluorescence intensity of microplastics in the residual solution and the original solution, respectively, and the number of sponges filled in the chromatographic filtration method in S2 can be 1, 2, 3, 4, 5, or 6.

[0029] Embodiment 3: Reference Figure 1-17 , which is basically the same as Example 1, the adsorption mechanism of sponge and sodium chloride in S3 was explored by repeated extrusion and adsorption of 50 mg sponge and 40 mL of 25 mg / L fluorescent microplastic solution (PS: 100 nm; PMMA: 5 μm) through kinetic experiments. Figure 8-11 As shown in the figure, the adsorption mechanism of the two was explored by repeatedly squeezing and adsorbing 50 mg of sponge with 40 mL of 25 mg / L fluorescent microplastic solution (PS: 100 nm; PMMA: 5 μm) through a kinetic experiment. The concentration of microplastics in the system at different time points (0, 1, 3, 6, 11, 17, 22 and 24 h) after the reaction was determined by a standard curve, as shown in the figure below. Fig.12 The adsorption amount of microplastics on the sponge is calculated as shown in formulas (3-1) and (3-2).

[0030] (3-1) (3-2) Among them, C 0 (mg L -1 ) and C e (mg L -1 ) represent the initial concentration and equilibrium concentration of fluorescent microplastics, respectively. C t (mg L -1 ) is the concentration of microplastics in the solution at a given time point. In addition, q e (mg g -1 ) and q t (mg g -1) represent the adsorption capacity at equilibrium and at a given time point, respectively. The mass of the sponge and the reaction volume are represented by m (g) and V (L), respectively. In S3: The fitting calculations of pseudo-first-order kinetics, pseudo-second-order kinetics, Elovich model and Webber-Morris model involved in the kinetics are shown in formulas (3-3), (3-4), (3-5) and (3-6).

[0031] (3-3) (3-4) (3-5) (3-6) Among them, k 1 (μg g -1 ), k 2 (μg g -1 ), k w (μg -1 h -0.5 h ) and k e is the corresponding adsorption rate constant; β is a parameter related to the activation energy and the adsorbent surface coverage of chemical adsorption.

[0032] The pH conditions in S3 were set to 2, 4, 7 and 11; the sodium chloride concentrations were set to 20 mM, 50 mM, 100 mM and 500 mM.

[0033] Furthermore, regarding the adsorption mechanism of microplastics on starch-gelatin sponge, fitting calculations revealed that the pseudo-second-order model and the Elovich model had the best fit, and it was found that it followed a physical adsorption process of heterogeneous diffusion.

[0034] Embodiment 4: Reference Figure 1-17 , which is basically the same as Example 1. In step S4, regarding the enzyme degradation determination step, a certain amount of sponge is taken, and the sponge after removing 5μm and 100nm is reacted in a system containing α-amylase (3,000 U) and saccharifying enzyme. Finally, the concentration of glucose in the system at different time points is determined. When soil degradation occurs, three sponges of different masses are selected and buried in the soil at a depth of 5cm, and the remaining weight of the sponge is weighed at different time points. The results of the degradation ability of microorganisms on sponges are shown in Fig.14 As shown in Figure 2, the ability of enzymes to decompose sponges is as follows: Fig.15 and 16 As shown in Figure 2, the soil environment affects the degradation of sponges. Fig.17 shown.

[0035] Furthermore, in the biocompatibility evaluation of S4, the mass fraction of the sponge was (0.05%, 1%, 2% and 6%, w / v), the reaction system contained 1 mL of Pichia. Pastoris in logarithmic growth phase per 30 mL of YPD medium, 1 mL of Escherichia coli in logarithmic growth phase per 30 mL of LB medium, and 1 mL of Lactic acid bacteria in logarithmic growth phase per 30 mL of MRS medium, and the reaction temperature was 37°C. The growth curve values ​​of the above three microorganisms are OD at 600 nm. 600 .

[0036] Furthermore, the amount of sponge used in the glucose determination of S4 was 200 mg, the degradation system was 20 mL of saline containing α-amylase (3,000 U) and saccharifying enzyme (5,000 U), the reaction temperature was 37°C, and unused starch / gelatin sponge was used as the control group. The determination time points were 0h, 4h, 6h, 9h, 16h, 21h and 24h.

[0037] Furthermore, the soil degradability evaluation in S4 was performed with initial sponge masses of 71 mg, 63.9 mg, and 60.6 mg, weighed at 0, 5, and 12 days, respectively.

[0038] It should be added that the reasons for selecting starch and gelatin as the matrix in the present invention are: Corn starch has a mature low-cost production process and a high content of amylopectin, which has a good foundation in production and application; amylopectin is rich in polyhydroxy structures and has a higher degree of cross-linking; in addition, the multi-scale structure, easy gelatinization and retrogradation characteristics of amylopectin make it easier to be fully utilized in the sponge preparation process. Gelatin has a wide range of sources, low cost, and high nutritional value. As a food raw material, there is no limit on the amount of use to ensure the safety of the raw material. Its own good biodegradability and biocompatibility have good recognition in meeting the characteristics of sponge environmental protection and no secondary pollution; in addition, gelatin is composed of a variety of amino acids connected to each other through carboxyl and amino groups, and functional groups such as amino, carboxyl and hydroxyl groups are more prone to self-crosslinking and cross-crosslinking. The complementary use of starch and gelatin not only makes up for the shortcomings of the weak strength of the starch skeleton after sponge shaping, but also improves the disadvantages of low ductility and brittle texture of gelatin after drying.

[0039] Explanation for choosing polystyrene and polymethyl methacrylate plastics as research objects: Polystyrene is a general-purpose plastic widely used in the manufacture of various consumer goods. As a hard solid plastic, it is usually used in food packaging, lightweight protective packaging, dining tableware, etc., and is closely related to food and human life. Polymethyl acrylate plastic is a representative plastic widely used in industrial production. For example, optical glass, TV screens, acrylic boards, product advertising windows, architectural glass, microfluidic chips, etc., are widely produced and used in industry. In addition, both have been reported to be detected in organisms, which are representative of hazards.

[0040] The above description is only a preferred specific implementation manner of the present invention, but the protection scope of the present invention is not limited thereto. Any technician familiar with the technical field can make equivalent replacements or changes according to the technical scheme and inventive concept of the present invention within the technical scope disclosed by the present invention, which should be covered by the protection scope of the present invention.

Claims

1. A preparation method for a plastic sponge of pullulan and gelatin and its application in removing microplastics, characterized in that: The following steps are involved: S1: Preparation of starch and gelatin semi-interpenetrating sponge: 4% starch and 3% gelatin were cross-linked, stirred evenly at an initial ratio of 1:1, and freeze-dried at -80℃ for 48h; S2: Adsorption treatment of starch-gelatin sponge and microplastics was performed. The sponge prepared after optimizing various conditions was squeezed and adsorbed with microplastic solutions in different food environments; S3: To investigate the state of the adsorption mechanism of microplastics and sponges, 50 mg of sponges were used to adsorb 5 mL of 25 mg / mL microplastic solutions at different pH and sodium chloride concentrations; S4: To evaluate the degradability of starch-gelatin sponges, different masses of sponges were cultured with corresponding bacterial colonies in different types of culture media of the same mass. The OD of the bacterial colonies at different time points was measured. 600 Determine the biocompatibility of the sponge.

2. The preparation of a plastic sponge of pullulan and gelatin and its application in microplastic removal according to claim 1, characterized in that: In step S1, the concentration of the starch solution is fixed at 4%, and the concentration of the starch solution is optimized under different preparation conditions.

3. The preparation of a plastic sponge of pullulan and gelatin and its application in microplastic removal according to claim 1, characterized in that: In step S1, the starch suspension is prepared by gelatinizing waxy corn starch at 92° C. (500 rpm) for 1 h.

4. The preparation of a plastic sponge of pullulan and gelatin and its application in microplastic removal according to claim 1, characterized in that: In step S1, the gelatin solution was chemically cross-linked by slowly adding formaldehyde (0.35%, v / v) to the above system under continuous stirring at 70°C within 1 minute.

5. The preparation of a plastic sponge of pullulan and gelatin and its application in microplastic removal according to claim 1, characterized in that: The operation method in step S2 is divided into a three-way valve extrusion method and a chromatography filtration method.

6. The preparation of a plastic sponge of pullulan and gelatin and its application in microplastic removal according to claim 1, characterized in that: In step S3, the adsorption mechanism of sponge and microplastics was investigated by repeatedly squeezing and adsorbing 50 mg of sponge with 40 mL of 25 mg / L fluorescent microplastic solution (PS: 100 nm; PMMA: 5 μm) through kinetic experiments.

7. The preparation of a plastic sponge of pullulan and gelatin and its application in microplastic removal according to claim 1, characterized in that: In step S4, regarding the enzyme degradation determination step, a certain amount of sponge is taken, and the sponge after removing 5 μm and 100 nm is reacted in a system containing α-amylase (3,000 U) and saccharifying enzyme. Finally, the concentration of glucose in the system at different time points is determined.

8. The preparation of a plastic sponge of pullulan and gelatin and its application in microplastic removal according to claim 1, characterized in that: Regarding the biocompatibility evaluation in step S4, the mass fraction of the sponge was (0.05%, 1%, 2% and 6%, w / v), the reaction system contained 1 mL of Pichia. Pastoris in the logarithmic growth phase per 30 mL of YPD medium, 1 mL of Escherichia coli in the logarithmic growth phase per 30 mL of LB medium, and 1 mL of Lactic acid bacteria in the logarithmic growth phase per 30 mL of MRS medium, and the reaction temperature was 37°C.

9. The preparation of a plastic sponge of pullulan and gelatin and its application in microplastic removal according to claim 1, characterized in that: In step S4, the amount of sponge used in the glucose determination was 200 mg, the degradation system was 20 mL of normal saline containing α-amylase (3,000 U) and saccharifying enzyme (5,000 U), and the reaction temperature was 37°C.

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