Micro-plastic poisoning antidote and application thereof in feed

Abstract

The invention discloses a microplastic poisoning antidote and application thereof in feed. The antidote comprises the following main components in percentage by mass: 25%-30% of hyocholic acid, 20%-30% of hyodeoxycholic acid, 20%-30% of chenodeoxycholic acid, 5%-15% of 7-ketolithocholic acid and 5%-15% of glycoursodeoxycholic acid. The antidote has positive and effective effects on microplastic poisoning, can significantly relieve microplastics, reduce the feed intake, daily gain and early feed conversion ratio of ducks, and also can significantly relieve the influence that microplastics induce liver immune cell infiltration. The experimental results prove that the antidote can effectively relieve the negative influence of microplastic toxicity on ducks.

Description

Technical Field

[0001] The present invention belongs to the technical field of plastic detoxification, and in particular relates to an antidote for microplastic poisoning and application thereof in feed. Background Art

[0002] Microplastics refer to plastic particles with a diameter of less than 5 mm, which are usually produced by the decomposition and wear of plastic products and waste in industrial processes. Due to their small size, these plastic particles can easily enter the environment, including water, soil and air, and accumulate in animals and humans through the food chain. Microplastics come from various sources, including microbeads in cosmetics, wear of synthetic fibers, and degradation of plastic products. They have a significant impact on the production performance of organisms, and their mechanisms and consequences have become a research hotspot with widespread concern.

[0003] Microplastics affect the production performance of organisms through a variety of mechanisms. First, microplastic particles entering the body of an organism may cause mechanical damage to the digestive tract, block the digestive system, and affect nutrient absorption and digestive function. Secondly, microplastics can absorb harmful chemicals in the environment, such as heavy metals, polychlorinated biphenyls, etc., and become carriers of pollutants. After organisms ingest these microplastics, harmful substances are released into the body, causing toxic effects. In addition, chemicals such as plasticizers and flame retardants contained in microplastics can interfere with the endocrine system of organisms, affect the normal function of hormones, and then affect growth, development and reproductive capacity. Finally, long-term exposure to microplastics may cause a stress response of the immune system and produce chronic inflammation, thereby affecting the health and production performance of organisms.

[0004] The impact of microplastics on organisms and ecosystems is multifaceted, and its main consequences include reduced biological health, impacts on food chains and ecological balance, economic losses, and increased risks to human health. The direct physical and chemical damage of microplastics to organisms can lead to deterioration of health conditions, manifested as slow growth, decreased immunity, and reduced reproductive capacity. In addition, the widespread presence of microplastics in the environment causes them to accumulate through the food chain, affecting organisms at different levels and ultimately threatening the balance of the ecosystem. For example, fish ingesting microplastics may affect predators, including birds and mammals. For humans, as part of the food chain, humans may ingest microplastics through their diet. These microplastics and the harmful substances they carry may pose a potential threat to human health, such as digestive system problems and endocrine disorders. For the economy, industries based on aquatic organisms, such as fisheries and aquaculture, will suffer from reduced production and quality due to microplastic pollution, leading to economic losses.

[0005] A large number of researchers have made unremitting efforts and explorations to reduce the toxic and side effects of microplastics, and have accumulated rich experience and methods. At present, the methods for reducing microplastics are mainly divided into two categories: degradation and adsorption. For example, the active ingredients of the composite microbial agents derived from Geobacillus thermodenitrificans and Brevibacillus thermoruber can promote the degradation of polyethylene microplastics. In addition, natural limestone particles can remove 70% of polystyrene, carboxyl-modified polystyrene and amino-modified polystyrene nanoplastic microspheres in water. Although the above substances have shown certain effects in reducing microplastics, their application in livestock and poultry still has limitations and needs further verification and optimization. Recent studies have shown that selenium-rich diets can significantly reduce the enrichment of microplastics in ephedra chickens. This discovery provides new ideas and methods for the removal of microplastics in livestock and poultry. At the same time, the study also found that the combination of licorice, isoliquiritigenin and glycyrrhetinic acid can alleviate the increase in serum CRP, TNF-α, IL-6 and IL-1β expression levels induced by microplastics, and reduce pathological damage to the intestines and liver. These natural ingredients have shown potential in alleviating the negative impact of microplastics on organisms.

[0006] In summary, although there has been some progress in the degradation and removal of microplastics, effective solutions for microplastics in livestock and poultry still need further research and verification. In recent years, food safety has been a focus of attention, and reducing microplastics in livestock and poultry meat, eggs and milk is also a hot topic. Therefore, it is particularly important to explore a green, efficient, safe, convenient and cost-effective natural scavenger to reduce residues in animal products in animal feed.

[0007] Bile acid is an important component of bile. Hepatocytes use cholesterol as a substrate and generate a class of amphipathic molecules with diverse structures under the action of a series of enzymes. These molecules are stored in the gallbladder and secreted into the duodenum under the action of cholecystokinin (CCK). About 95% of bile acids in the intestine return to the liver by reabsorption. In recent years, a large number of studies have shown that bile acids, as signaling molecules, play an important role in regulating the body's glucose and lipid metabolism, antibacterial and anti-inflammatory, and detoxification by activating nuclear receptors α (such as liver X receptor α) or other ligand-dependent transcription factors (such as retinoic acid X receptor α).

[0008] At present, there is no relevant literature reporting on the role of bile acid in reducing microplastic residues in ducks. Summary of the invention

[0009] The purpose of the present invention is to provide a green, efficient, safe, convenient and cost-effective natural mitigator, which is non-toxic, pollution-free and drug-resistant, and can significantly reduce the toxic and side effects of microplastics on the body, in view of the fact that existing detoxifiers are still limited or unsuitable for removing microplastics in livestock and poultry.

[0010] The purpose of the present invention is achieved through the following technical solutions:

[0011] An antidote for microplastic poisoning, comprising the following main components and their mass percentages: 25% to 30% of hyocholic acid, 20% to 30% of hyodeoxycholic acid, 20% to 30% of chenodeoxycholic acid, 5% to 15% of 7-ketolithocholic acid and 5% to 15% of glycoursodeoxycholic acid.

[0012] Preferably, the total bile acid content in the antidote is ≥ 95%.

[0013] More preferably, the antidote comprises the following main components and their mass percentages: 28.95% hyodocholic acid, 24.25% hyodeoxycholic acid, 25.71% chenodeoxycholic acid, 10.26% 7-ketolithocholic acid and 9.60% glycoursodeoxycholic acid, and the total bile acid content is 98.77%.

[0014] The antidote for microplastic poisoning is a mixed bile acid prepared from pig bile paste powder as a raw material, which is subjected to saponification, oxidative decolorization, acidification, water washing to remove water-soluble impurities, recrystallization, and drying.

[0015] Another object of the present invention is to provide a method for preparing the antidote for microplastic poisoning, comprising the following steps:

[0016] (1) Saponification: The crushed pig bile paste powder and 10% sodium hydroxide solution by mass fraction are mixed at a solid-liquid ratio of 1:10 kg / L, heated to 100° C., and kept boiling for 12 hours; after the reaction is completed, the saponified solution is cooled until solid-liquid separation occurs, the supernatant is removed, and the solid is the bile paste saponified product;

[0017] (2) Oxidative decolorization: Mix the bile paste saponified product and water, heat to 80°C to completely dissolve the bile paste saponified product, add hydrogen peroxide, mix well, react at room temperature for 24 hours, filter the reaction liquid to obtain a filtrate; wherein the mass ratio of the bile paste saponified product to water is 1:6; the solid-liquid ratio of the bile paste saponified product to hydrogen peroxide is 1:0.3 kg / L;

[0018] (3) Acidification: Under stirring, slowly add 10% by mass hydrochloric acid solution to adjust the pH value of the filtrate to 3 to obtain a suspension containing white solids;

[0019] (4) Purification: The suspension is centrifuged, and water is added for rinsing after each centrifugation until the supernatant is neutral; the white solid obtained by centrifugation is dried to obtain crude bile acid;

[0020] (5) Recrystallization: Mix the crude bile acid with ethyl acetate at a solid-liquid ratio of 1:10 kg / L, stir until the crude bile acid is dissolved, add anhydrous sodium sulfate for dehydration, filter, and collect the filtrate; concentrate the filtrate under reduced pressure to obtain bile acid;

[0021] (6) Drying: Drying the bile acid to obtain a mixed bile acid product.

[0022] Another object of the present invention is to provide the use of the antidote in the preparation of feed for preventing microplastic poisoning.

[0023] Another object of the present invention is to provide a feed for preventing microplastic poisoning, which contains the antidote described in the present invention, and the addition ratio of the antidote is 150 to 400 mg of antidote per 1000 g of feed.

[0024] Preferably, the antidote is added in an amount of 150 mg, 250 mg, 300 mg or 400 mg per 1000 g of feed.

[0025] Most preferably, the antidote is added in an amount of 250 mg of antidote per 1000 g of feed.

[0026] Preferably, the feed for preventing microplastic poisoning is duck feed for preventing microplastic poisoning.

[0027] Specifically, the duck breed is Zhijiang duck.

[0028] Another object of the present invention is to provide a method for preparing feed for preventing microplastic poisoning, comprising the step of mixing the antidote described in the present invention into the feed.

[0029] The present invention has the following beneficial effects:

[0030] Compared with other common protective agents, the antidote for microplastics of the present invention is a natural extract, which has the characteristics of stable and reliable source, high cost performance, convenient use and significant effect. The antidote of the present invention does not contain any antibiotics, has no toxic side effects, no drug resistance and no environmental pollution, and is a green and efficient antidote.

[0031] Experiments have shown that the antidote has a positive and effective effect on microplastic poisoning. It can significantly alleviate the reduction of ducks' feed intake, daily weight gain and early feed-to-meat ratio caused by microplastics, and also alleviate microplastic-induced liver pathological changes. BRIEF DESCRIPTION OF THE DRAWINGS

[0032] Figure 1 The average daily feed intake (ADFI) of ducks in different treatment groups in two stages.

[0033] Figure 2 The average daily gain (ADG) of ducks in different treatment groups in two stages.

[0034] Figure 3 This is the feed-to-meat ratio (FCR) of ducks in different treatment groups in two stages.

[0035] Figure 4 Hematoxylin-eosin stained tissue sections of duck livers in different treatment groups. DETAILED DESCRIPTION

[0036] The technical solution of the present invention is further described below in conjunction with the implementation, but the implementation of the present invention is not limited thereto. The test methods used in the following implementation are all conventional methods unless otherwise specified.

[0037] Example 1

[0038] The antidote for preventing and treating microplastic poisoning is a mixed bile acid made from pig bile paste powder. The preparation steps are as follows:

[0039] (1) Saponification: Weigh 200 kg of crushed pig bile paste powder and add it to a reactor. Add 2000 L of 10% sodium hydroxide solution at a solid-liquid ratio of 1:10 kg / L. When the solution in the reactor is heated to 100°C, start timing and keep boiling for 12 hours. After the reaction is completed, cool the saponified solution until solid and liquid layers are separated. Use a pipeline centrifugal pump to pump the supernatant into a storage tank. The remaining solid is the bile paste saponified product. Separate the protein, lipid and other substances in the bile paste through saponification.

[0040] (2) Oxidative decolorization: Add 1200 kg of tap water to the reactor and heat to 80°C to completely dissolve the bile paste saponified product (the mass ratio of tap water to pig bile paste powder is 6:1); use a pipeline centrifugal pump to pump the solution in the reactor into the decolorization tank, add 60 L of hydrogen peroxide, mix evenly, and react at room temperature for 24 hours; after the reaction is completed, filter the solution into an acidification tank; separate the pigment and residual protein through oxidative decolorization.

[0041] (3) Acidification: Slowly add 10% by mass hydrochloric acid solution into the acidification tank, stirring continuously. When the pH value of the solution is about 3, the sodium salt is reduced to crystalline bile acid. Stop adding acid, and a suspension containing white solids is obtained.

[0042] (4) Purification, washing with water to remove water-soluble impurities: The suspension is centrifuged, and water is added for washing after each centrifugation. When the centrifuged water is detected to be neutral, the washing is completed. The water-soluble impurities attached to the surface of the bile acid solid are removed by washing with water; the white solid obtained by centrifugation is placed in an oven and dried at 100°C until the moisture content is no more than 10%, thereby obtaining a crude bile acid product.

[0043] (5) Recrystallization: Put the crude bile acid into an extraction tank, add ethyl acetate (the mass volume ratio of the crude bile acid to ethyl acetate is 1:10 kg / L), stir and dissolve, and when the crude bile acid is basically dissolved, add anhydrous sodium sulfate for dehydration, filter, and collect the filtrate; the filtrate is sent to a concentration tank for reduced pressure concentration, and the ethyl acetate is recovered to obtain bile acid.

[0044] (6) Drying: Spread the bile acid on a stainless steel drying tray, put it into an oven, introduce steam, and dry it at 100° C. When the bile acid becomes a white powder, take a sample to detect the moisture content to be no more than 1%, thereby obtaining a mixed bile acid product.

[0045] The mixed bile acid finished product mainly contains 28.95% of hyodocholic acid, 24.25% of hyodeoxycholic acid, 25.71% of chenodeoxycholic acid, 10.26% of 7-ketolithocholic acid and 9.60% of glycoursodeoxycholic acid, with a total content of 98.77%.

[0046] Example 2

[0047] Experiment on mitigation of the toxic side effects of microplastics in Zhijiang ducks by antidotes:

[0048] 500 μg of microplastics were added to 1000 g of feed to prepare a diet containing 500 μg of microplastics per kilogram of feed.

[0049] Take 1000 g of the diet containing 500 μg of microplastics, add 250 mg of the antidote of Example 1 and mix well.

[0050] Experimental method: One-day-old Zhijiang ducks (half male and half female) were selected and raised according to the standard feeding process. On the 3rd day, they were randomly divided into 3 groups: a negative control group (CON group, without adding microplastics and antidotes), a positive group (microplastic group, i.e., PLC group, with microplastics but no antidote), and an experimental group (microplastic + BA group, i.e., PLB group, with microplastics and antidotes added). There were 15 ducks in each group, and there was no significant difference in the initial weight of the ducks among the groups (p>0.05). The feeding method of each group is as follows: PLB group: from the 4th day, the diet containing microplastics and bile acid was fed every day, the dose of microplastics in the diet was 500μg / kg feed, and the dose of antidote was 250mg / kg feed; PLC group: the diet without antidote but microplastics was fed, from the 4th day, the diet containing microplastics was fed every day, and the dose of microplastics in the diet was 500μg / kg feed; CON group: the diet without microplastics and antidote was fed. During the experiment, free drinking water and feeding were allowed, feces were cleaned daily, disinfected regularly, and temperature and humidity were controlled according to the standard paradigm. The experimental period was 15 days. The feed intake was collected every day, weighed on the 7th and 15th days and used to calculate the corresponding indicators. After the experiment, each group fasted for 9h and weighed on an empty stomach. Blood was taken by decapitation, and the liver was taken in a standardized manner and placed in a 4% polyformaldehyde solution for subsequent slice analysis.

[0051] Depend on Figure 1 It can be seen that compared with the CON group, the average daily feed intake (ADFI) of ducks in the PLC group at two different stages was significantly reduced (p < 0.05); compared with the PLC group, the antidote (PLB group) significantly increased the ADFI of ducks at two different stages (p < 0.001). This shows that the antidote of the present invention can significantly increase the feed intake of ducks.

[0052] Depend on Figure 2 It can be seen that compared with the CON group, the average daily weight gain (ADG) of ducks in the PLC group at two different stages was significantly reduced (p < 0.05); compared with the PLC group, the antidote (PLB group) significantly increased the ADG of ducks at two different stages (p < 0.01), indicating that the antidote of the present invention can significantly increase body weight.

[0053] Depend on Figure 3 It can be seen that compared with the CON group, the feed-to-meat ratio (FCR) of the PLC group increased at two different stages, and there was a significant difference in the first stage (p < 0.05); compared with the PLC group, the antidote (PLB group) tended to reduce the FCR of the first stage (p = 0.08).

[0054] Depend on Figure 4 It can be seen that compared with the CON group, the infiltration of immune cells in the liver of 19-day-old ducks in the PLC group was significantly increased, and compared with the PLC group, the antidote (PLB group) significantly reduced the infiltration of immune cells in the liver of ducks.

[0055] In summary, after ducks ingested microplastics, their growth performance was significantly reduced. However, adding the antidote of the present invention to the diet could alleviate the effects of microplastics on the ducks' feed intake and conversion efficiency, and could also significantly alleviate the effects of microplastics inducing liver immune cell infiltration. This confirms that the antidote of the present invention can effectively alleviate the negative effects of microplastic toxicity on Zhijiang ducks.

[0056] The embodiments described above are only descriptions of the preferred implementation modes of the present invention, and are not intended to limit the scope of the present invention. Without departing from the design spirit of the present invention, various modifications and improvements made to the technical solutions of the present invention by ordinary engineering and technical personnel in the field should fall within the protection scope determined by the claims of the present invention.

Claims

1. An antidote for microplastic poisoning, characterized in that: The antidote comprises the following main components and their mass percentages: 25% to 30% of hyocholic acid, 20% to 30% of hyodeoxycholic acid, 20% to 30% of chenodeoxycholic acid, 5% to 15% of 7-ketolithocholic acid and 5% to 15% of glycoursodeoxycholic acid.

2. The antidote for microplastic poisoning according to claim 1, characterized in that: In the antidote, the total bile acid content is ≥95%.

3. The antidote for microplastic poisoning according to claim 1, characterized in that: The antidote comprises the following main components and their mass percentages: 28.95% of hyocholic acid, 24.25% of hyodeoxycholic acid, 25.71% of chenodeoxycholic acid, 10.26% of 7-ketolithocholic acid and 9.60% of glycoursodeoxycholic acid, and the total bile acid content is 98.77%.

4. A method for preparing an antidote for microplastic poisoning according to any one of claims 1 to 3, characterized in that: The following steps are involved: The invention takes pig gallbladder paste powder as raw material and is prepared through the steps of saponification, oxidation decolorization, acidification, water washing to remove water-soluble impurities, recrystallization and drying.

5. The preparation method according to claim 4, characterized in that: The following steps are involved: (1) Saponification: The crushed pig bile paste powder and 10% sodium hydroxide solution by mass fraction are mixed at a solid-liquid ratio of 1:10 kg / L, heated to 100° C., and kept boiling for 12 hours; after the reaction is completed, the saponified solution is cooled until solid-liquid separation occurs, the supernatant is removed, and the solid is the bile paste saponified product; (2) Oxidative decolorization: Mix the bile paste saponified product and water, heat to 80°C to completely dissolve the bile paste saponified product, add hydrogen peroxide, mix well, react at room temperature for 24 hours, filter the reaction liquid to obtain a filtrate; wherein the mass ratio of the bile paste saponified product to water is 1:6; the solid-liquid ratio of the bile paste saponified product to hydrogen peroxide is 1:0.3 kg / L; (3) Acidification: Under stirring, slowly add 10% by mass hydrochloric acid solution to adjust the pH value of the filtrate to 3 to obtain a suspension containing white solids; (4) Purification: The suspension is centrifuged, and water is added for rinsing after each centrifugation until the supernatant is neutral; the white solid obtained by centrifugation is dried to obtain crude bile acid; (5) Recrystallization: Mix the crude bile acid with ethyl acetate at a solid-liquid ratio of 1:10 kg / L, stir until the crude bile acid is dissolved, add anhydrous sodium sulfate for dehydration, filter, and collect the filtrate; concentrate the filtrate under reduced pressure to obtain bile acid; (6) Drying: Drying the bile acid to obtain a mixed bile acid product.

6. Use of the antidote described in any one of claims 1 to 3 in the preparation of feed for preventing microplastic poisoning.

7. A feed for preventing microplastic poisoning, characterized in that: The feed contains the antidote according to any one of claims 1 to 3.

8. The feed for preventing microplastic poisoning according to claim 7, characterized in that: The adding ratio of the antidote is 150-400 mg of the antidote per 1000 g of feed.

9. The feed for preventing microplastic poisoning according to claim 8, characterized in that: The adding ratio of the antidote is 250 mg of the antidote per 1000 g of feed.

10. The feed for preventing microplastic poisoning according to claim 7, characterized in that: The feed is duck feed for preventing microplastic poisoning.

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