A mycotoxin attenuator for improving mitochondrial function and application thereof

Abstract

The application discloses a mycotoxin attenuant for improving mitochondrial function and application, and has the following components: 10-20 parts by weight of L-malic acid, 3-5 parts by weight of alpha-ketoglutaric acid (salt), 15-25 parts by weight of itaconic acid and 1-3 parts by weight of 5-norwongenin. In mycotoxin contaminated feed, the addition of L-malic acid, alpha-ketoglutaric acid (salt), itaconic acid or 5-norwongenin alone cannot improve mitochondrial function or relieve mycotoxin toxicity. Compared with the combination of any one of the four components, the complete addition of the four components can significantly enhance mitochondrial oxidative phosphorylation, weaken glycolysis, obviously improve mitochondrial energy metabolism, effectively relieve mycotoxin-induced mitochondrial damage in the intestines and liver of livestock and poultry, and thus improve the production performance and health status of livestock and poultry. The application can significantly reduce the toxic effect of mycotoxin on livestock and poultry through the synergistic effect of the components.

Description

Technical Field

[0001] This invention relates to the field of additives for livestock and poultry farming, specifically to a mycotoxin attenuator that improves mitochondrial function and its application. Background Technology

[0002] Currently, approximately 400 mycotoxins have been identified, of which more than 350 can be detected in feed. Aflatoxin, vomitoxin (deoxynivalenol), zearalenone, ochratoxin, fumonisin, and T-2 toxin are common mycotoxins in feed. Even if the mycotoxin contamination level in feed is within the relevant feed hygiene standards, the synergistic effect of multiple toxins still harms livestock health and production performance. Therefore, mycotoxin attenuation technology in feed is of paramount importance.

[0003] The reduction of mycotoxins in feed mainly occurs through adsorption and degradation. Most adsorbents can only adsorb highly polar mycotoxins such as aflatoxin, while their adsorption effect on toxins such as vomitoxin is poor. Problems with the biodegradation of mycotoxins include: the tendency of detoxifying strains to degenerate, unstable degradation activity, and incompatibility between growth conditions and the gastrointestinal environment of livestock and poultry; the poor stability of detoxifying enzymes, and the high specificity of degrading enzymes, coupled with the presence of multiple mycotoxins in feed, making it difficult for a single enzyme to comprehensively cover all types, thus requiring the formulation of multiple enzymes.

[0004] Mitochondria are the targets of mycotoxins and a key hub for toxic signal transduction. The core mechanisms of mycotoxin-induced intestinal damage, hepatotoxicity, and inflammation are all closely related to mitochondrial dysfunction. Targeting mitochondria to antagonize the toxic effects of mycotoxins has the advantage of universality: regardless of the type and source of the mycotoxin, one of its core toxic mechanisms is the destruction of mitochondrial function. Therefore, nutritional regulation aimed at protecting mitochondria and enhancing their function is a universal strategy for intercepting toxic pathways at the "end point," and can address the synergistic attack of various mixed toxins.

[0005] In summary, although existing adsorption and biodegradation technologies have made some progress in mycotoxin attenuation, they all suffer from limitations such as narrow adsorption spectra, unstable degradation activity, and the inability to cope with the synergistic contamination of multiple toxins due to the limited number of enzymes used. Mitochondria, as the core target of multiple mycotoxins and a key hub for toxic signal transduction, offer the advantage of universality in strategies that protect their functional integrity, transcending toxin types and independent of toxin structure recognition. Therefore, there is an urgent need to develop a mycotoxin attenuator that focuses on improving mitochondrial function from a nutritional regulation perspective, thereby effectively blocking the synergistic toxic effects of different toxins at the mitochondrial level. Summary of the Invention

[0006] The purpose of this invention is to overcome the shortcomings of conventional adsorption and degradation technologies in the field of mycotoxin attenuation, and to provide a mycotoxin attenuator that improves mitochondrial function.

[0007] To achieve the above-mentioned objectives, the technical solution adopted by this invention is as follows:

[0008] In a first aspect, the present invention provides a mycotoxin attenuator that improves mitochondrial function, comprising the following components in parts by weight: 10-20 parts of L-malic acid, 3-5 parts of α-ketoglutarate or α-ketoglutarate salt, 15-25 parts of itaconic acid, and 1-3 parts of 5-norhesperidin.

[0009] It should be noted that the 5-demethylnobiletin (CAS No.: 2174-59-6) used in this invention is a polymethoxyflavonoid compound found in citrus plants, which can be prepared by existing methods or purchased through commercial channels.

[0010] As a preferred embodiment of the first aspect above, the mycotoxin attenuator is composed of 10-20 parts of L-malic acid, 3-5 parts of α-ketoglutarate or α-ketoglutarate, 15-25 parts of itaconic acid, and 1-3 parts of 5-norhesperidin, by weight.

[0011] As a preferred embodiment of the first aspect above, the α-ketoglutarate is sodium α-ketoglutarate or calcium α-ketoglutarate.

[0012] In a second aspect, the present invention provides a feed containing a mycotoxin attenuator that improves mitochondrial function as described in any of the first aspects above.

[0013] As a preferred embodiment of the second aspect above, the amount of mycotoxin attenuator added to the feed is 0.04 wt.% to 0.05 wt.%.

[0014] As a preferred embodiment of the second aspect above, the feed is pig feed or broiler feed.

[0015] Thirdly, the present invention provides the application of a mycotoxin attenuator that improves mitochondrial function as described in any of the first aspects above in livestock and poultry feeding, specifically for alleviating mycotoxin-induced mitochondrial damage in the intestines and / or liver of livestock and poultry.

[0016] As a preferred embodiment of the third aspect above, the mycotoxin attenuator that improves mitochondrial function is added to the feed for livestock and poultry in the form of an additive.

[0017] As a preferred embodiment of the third aspect above, the mitigation of mitochondrial damage is manifested in enhanced mitochondrial oxidative phosphorylation, reduced glycolysis, and improved mitochondrial energy metabolism.

[0018] As a preferred embodiment of the third aspect above, the livestock or poultry is pig or broiler chicken.

[0019] The present invention can also provide a feed containing the above-mentioned mycotoxin attenuator. The amount of mycotoxin attenuator added to the feed is 0.04 wt.%~0.05 wt.%.

[0020] The mycotoxin attenuator of the present invention has the following characteristics:

[0021] (1) This invention identifies characteristic metabolic markers and key metabolic pathways of mycotoxin-induced mitochondrial damage through multi-omics analysis such as metabolomics and transcriptomics, and invents the mycotoxin attenuator that improves mitochondrial function through network pharmacology, high-throughput screening, molecular docking and other technical means.

[0022] (2) There are currently no reports on the combined use of L-malic acid, α-ketoglutarate (salt), itaconic acid, and 5-norhesperidin to alleviate mycotoxin toxicity in animals. Studies on their individual application in livestock and poultry have reported that L-malic acid alone requires 0.5% to promote growth, α-ketoglutarate requires 2%, and itaconic acid requires 1%. In this invention, L-malic acid is added at only 0.01~0.02%, α-ketoglutarate at 0.004~0.005%, and itaconic acid at 0.017~0.026%, all far below the effective doses of a single component. However, the combined use of the four components produces a highly significant synergistic effect, indicating that this invention is not a simple additive process, but rather activates the mitochondrial repair mechanism through multiple pathways, which is unforeseen by those skilled in the art. This demonstrates a significant non-obviousness.

[0023] (3) In mycotoxin-contaminated feed, the addition of L-malic acid, α-ketoglutarate (salt), itaconic acid, or 5-norhesperidin alone cannot improve mitochondrial energy metabolism or alleviate the toxic effects of mycotoxins. Compared with the four three-component combinations (i.e., combinations lacking one of each component), the addition of the feed mycotoxin attenuator of this invention can significantly enhance mitochondrial oxidative phosphorylation, reduce glycolysis, significantly improve mitochondrial energy metabolism, and effectively alleviate mycotoxin-induced mitochondrial damage in the intestines and liver of livestock and poultry, thereby improving livestock and poultry production performance and health status.

[0024] (4) L-malic acid, α-ketoglutarate (salt), itaconic acid, and 5-norhesperidin have a very significant synergistic effect in improving mitochondrial function and alleviating mycotoxin toxicity. The mycotoxin attenuator of the present invention can significantly alleviate the toxic effects of mycotoxins in feed on livestock and poultry through the synergistic effect of each component, and improve the production performance and health status of livestock and poultry. Attached Figure Description

[0025] Figure 1 The growth performance of Jinhua pigs in each group in Example 1 is shown, where different letters indicate significant differences (P<0.05).

[0026] Figure 2 The intestinal barrier, liver function, oxidative stress, and inflammatory markers of pigs in each group in Example 1 are represented by different letters, indicating significant differences (P<0.05).

[0027] Figure 3 The values ​​represent the mitochondrial energy metabolism and function indicators in each group of pigs in Example 1, with different letters indicating significant differences (P<0.05). The mean value of the control group was 1.00, and the values ​​of other groups were relative multiples.

[0028] Figure 4 The values ​​represent the growth performance, intestinal barrier, liver function, oxidative stress, and inflammation of pigs in each group in Example 2. Different letters indicate significant differences (P<0.05).

[0029] Figure 5 The values ​​represent the mitochondrial energy metabolism and function indicators for each group in Example 2, with different letters indicating significant differences (P<0.05). The mean value for the control group was 1.00, and the values ​​for other groups represent relative folds.

[0030] Figure 6 The values ​​represent the growth performance, intestinal barrier function, liver function, oxidative stress, and inflammatory markers of broilers in Example 3, with different letters indicating significant differences (P<0.05).

[0031] Figure 7 The values ​​represent the mitochondrial energy metabolism and function indicators of each group of broilers in Example 3, with different letters indicating significant differences (P<0.05). The mean value of the control group was 1.00, and the values ​​of other groups represent relative folds. Detailed Implementation

[0032] To make the above-mentioned objects, features, and advantages of the present invention more apparent and understandable, specific embodiments of the present invention will be described in detail below with reference to the accompanying drawings. Many specific details are set forth in the following description to provide a thorough understanding of the present invention. However, the present invention can be practiced in many other ways different from those described herein, and those skilled in the art can make similar modifications without departing from the spirit of the present invention. Therefore, the present invention is not limited to the specific embodiments disclosed below. Technical features in various embodiments of the present invention can be combined accordingly without mutual conflict.

[0033] This invention, through in-depth research into the pathological and molecular mechanisms of mycotoxins, reveals that mycotoxins damage the intestines and liver of livestock and poultry, significantly increase mitochondrial ROS production, cause mitochondrial swelling, vacuolation, cristae damage, and reduce mitochondrial membrane potential and respiratory chain complex activity, leading to mitochondrial dysfunction. Furthermore, mycotoxins weaken mitochondrial oxidative phosphorylation and enhance mitochondrial glycolysis, indicating a reprogramming of mitochondrial energy metabolism. Based on this, this invention provides a mycotoxin attenuator to improve mitochondrial function, comprising at least four components in parts by weight: 10-20 parts L-malic acid, 3-5 parts α-ketoglutarate or α-ketoglutarate salt, 15-25 parts itaconic acid, and 1-3 parts 5-norhesperidin. The α-ketoglutarate salt can be sodium α-ketoglutarate or calcium α-ketoglutarate.

[0034] It should be noted that the above four components are the core components of this mycotoxin attenuator. In theory, in addition to the above four components, other auxiliary components can be added to this mycotoxin attenuator, and there are no restrictions on this.

[0035] The mycotoxin attenuator for improving mitochondrial function described above in this invention can be added to a basic feed to form a feed with mycotoxin attenuation function. The recommended addition amount of the mycotoxin attenuator in the feed is 0.04 wt.% to 0.05 wt.%, and the basic feed can be pig feed or broiler feed, etc., depending on the specific type of livestock or poultry being targeted.

[0036] The aforementioned mycotoxin attenuators that improve mitochondrial function can be added to the feed of livestock and poultry such as pigs or broilers as additives during the breeding process. They are used to alleviate mycotoxin-induced mitochondrial damage in the intestines and / or liver of livestock and poultry. The alleviation of mitochondrial damage is manifested by enhancing mitochondrial oxidative phosphorylation, reducing glycolysis, and improving mitochondrial energy metabolism.

[0037] In the above-mentioned formulation, the four components were identified through multi-omics analysis, including metabolomics and transcriptomics, which determined the characteristic metabolic markers and key metabolic pathways of mycotoxin-induced mitochondrial damage. These components were further screened using techniques such as network pharmacology, high-throughput screening, and molecular docking. L-malic acid, α-ketoglutarate (salt), and itaconic acid have certain applications in livestock and poultry. However, when applying these components, L-malic acid alone needs to be added at 0.5% to promote growth, α-ketoglutarate at 2%, and itaconic acid at 1%. 5-Demethylnobiletin (CAS No.: 2174-59-6) is a polymethoxyflavonoid compound found in citrus plants and currently has some medicinal uses. There are currently no reports of combining L-malic acid, α-ketoglutarate (salt), itaconic acid, and 5-demethylnobiletin for mycotoxin detoxification in feed. Through experiments, this invention has found that L-malic acid, α-ketoglutarate (salt), itaconic acid, and 5-norhesperidin have a very significant synergistic effect in improving mitochondrial function and alleviating mycotoxin toxicity. They can significantly enhance mitochondrial oxidative phosphorylation, reduce glycolysis, significantly improve mitochondrial energy metabolism, and effectively alleviate mycotoxin-induced mitochondrial damage in the intestines and liver of livestock and poultry, thereby improving livestock and poultry production performance and health status.

[0038] The following examples demonstrate the specific application effects of the aforementioned mycotoxin attenuator. The materials used in each of the following examples can be commercially available products or prepared using methods reported in existing technology, and are not limited thereto.

[0039] Example 1

[0040] In this embodiment, 20 kg of L-malic acid, 5 kg of sodium α-ketoglutarate, 25 kg of itaconic acid, and 2 kg of 5-norhesperidin were mixed thoroughly to obtain a mycotoxin detoxifier for improving mitochondrial function (MDIM). The addition amount in Jinhua pig feed was 0.05 wt.%. For ease of subsequent description, this experimental group is referred to as the MDIM group.

[0041] One hundred and twenty Jinhua pigs with an average weight of 58 kg were randomly divided into ten groups: ① Control group (basal diet contaminated with mycotoxins); ② L-malic acid group; ③ Sodium α-ketoglutarate group; ④ Itaconic acid group; ⑤ 5-norhesperidin group; ⑥ Sodium α-ketoglutarate + itaconic acid + 5-norhesperidin group; ⑦ L-malic acid + itaconic acid + 5-norhesperidin group; ⑧ L-malic acid + sodium α-ketoglutarate + 5-norhesperidin group; ⑨ L-malic acid + sodium α-ketoglutarate + itaconic acid group; ⑩ MDIM group. The specific procedures for each group are as follows:

[0042] ① Control group: Feed contaminated with mycotoxins, the contents of aflatoxin B1, vomitoxin, zearalenone and fumonisin in the feed were 11, 907, 155 and 3062 μg / kg, respectively;

[0043] ②L-malic acid group: Based on the control group, L-malic acid was added alone to the same mycotoxin-contaminated feed;

[0044] ③Sodium α-ketoglutarate group: In addition to the control group, sodium α-ketoglutarate was added alone to the same mycotoxin-contaminated feed;

[0045] ④ Itaconic acid group: In addition to the control group, itaconic acid was added alone to the same mycotoxin-contaminated feed;

[0046] ⑤ The 5-norhesperidin group was supplemented with 5-norhesperidin alone in the same mycotoxin-contaminated feed, in addition to the control group.

[0047] ⑥ Sodium α-ketoglutarate + itaconic acid + 5-norhesperidin group: Based on the control group, sodium α-ketoglutarate, itaconic acid, and 5-norhesperidin were added to the same mycotoxin-contaminated feed.

[0048] ⑦ L-malic acid + itaconic acid + 5-norhesperidin group: Based on the control group, L-malic acid, itaconic acid and 5-norhesperidin were added to the same mycotoxin-contaminated feed.

[0049] ⑧L-malic acid + sodium α-ketoglutarate + 5-norhesperidin group: Based on the control group, L-malic acid, sodium α-ketoglutarate, and 5-norhesperidin were added to the same mycotoxin-contaminated feed.

[0050] ⑨ L-malic acid + sodium α-ketoglutarate + itaconic acid group: Based on the control group, L-malic acid, sodium α-ketoglutarate, and itaconic acid were added to the same mycotoxin-contaminated feed.

[0051] ⑩ MDIM group: This refers to the addition of the aforementioned composition MDIM in this embodiment, which contains all four components, with an MDIM addition amount of 0.05 wt.% in the feed.

[0052] Except for the control group and the MDIM group, the content of each component added in groups ② to ⑨ above is consistent with the content of the corresponding component in the feed of the MDIM group.

[0053] For the ten groups mentioned above, each group had three replicates, with four Jinhua pigs per replicate. The experimental period was 49 days, and the average daily weight gain and feed conversion ratio were calculated. After the feeding trial, six pigs from each group were slaughtered, and intestinal, liver, and blood samples were collected for testing. The results of each indicator are as follows: Figure 1 , Figure 2 and Figure 3 As shown.

[0054] Depend on Figure 1 As can be seen, compared with the control group, there were no significant differences in daily weight gain and feed conversion ratio in the single-drug groups ②~⑤ and the groups with any three components ⑥~⑨ (P>0.05), indicating that neither single nor three-component combinations could effectively alleviate growth inhibition caused by mycotoxins. However, when the four components were used in combination (MDIM group), daily weight gain was significantly increased and feed conversion ratio was significantly decreased (P<0.05). Bliss independence model analysis showed that the four components had a very strong synergistic effect, far exceeding the simple sum of the effects of the individual drugs. In summary, L-malic acid, sodium α-ketoglutarate, itaconic acid, and 5-norhesperidin, in a specific ratio, have a significant synergistic effect on alleviating growth inhibition caused by mycotoxins, producing unexpected technical effects in both daily weight gain and feed conversion ratio, two independent indicators.

[0055] Depend on Figure 2 As can be seen, compared with the control group, there were no significant differences in villous height / crypt depth (villous-crypt ratio), serum diamine oxidase (DAO), serum aspartate aminotransferase (AST) and alanine aminotransferase (ALT), serum total antioxidant capacity (TAC) and malondialdehyde (MDA), and serum TNF-α in the single-drug groups ②~⑤ and the groups with any three components ⑥~⑨ (P>0.05). This indicates that single or three-component combinations cannot effectively alleviate mycotoxin-induced intestinal and liver damage, oxidative stress, and inflammation. However, when the four components were used in combination (MDIM group), all of the above indicators were significantly improved (P<0.05). Bliss independence model analysis showed that the four components had a very strong synergistic effect, which far exceeded the simple sum of the effects of the individual drugs. In summary, L-malic acid, sodium α-ketoglutarate, itaconic acid, and 5-norhesperidin, in a specific ratio, have a significant synergistic effect on alleviating mycotoxin-induced intestinal and liver damage, oxidative stress, and inflammation, producing unexpected technical effects.

[0056] Depend on Figure 3As can be seen, compared with the control group, there were no significant differences in mitochondrial membrane potential (ΔΨm), mitochondrial ROS, respiratory chain complex I activity, cellular oxygen consumption rate (OCR), basal respiration (BR), maximal respiration (MR), ATP production, expression of the glycolysis-related gene PFKFB3, extracellular acidification rate (ECAR), and intestinal mucosal lactate content in each single-drug group ②~⑤ and any three-component combination group ⑥~⑨. Furthermore, morphological changes in intestinal mucosal and liver mitochondria (such as swelling, vacuolation, and cristae damage) were observed in all treatment groups (①-⑨). This indicates that neither a single component nor a combination of three components can effectively alleviate mitochondrial energy metabolism disorders and functional damage caused by mycotoxins. However, when the four components were used in combination (⑩MDIM group), all the above indicators were significantly improved (P<0.05), and no morphological changes in mitochondrial damage were observed in the intestinal mucosa and liver. Bliss independence model analysis showed that the four components had a very strong synergistic effect, far exceeding the simple summation of the effects of each single drug. In summary, L-malic acid, sodium α-ketoglutarate, itaconic acid, and 5-norhesperidin, when combined in specific ratios, exhibit significant synergistic effects on mycotoxin-induced mitochondrial energy metabolism disorders and functional impairment. They can significantly enhance mitochondrial oxidative phosphorylation, reduce glycolysis, improve mitochondrial energy metabolism, and alleviate intestinal and liver mitochondrial damage, producing unexpected technical effects.

[0057] Example 2

[0058] In this embodiment, 10 kg of L-malic acid, 3 kg of α-ketoglutarate, 20 kg of itaconic acid, and 1 kg of 5-norhesperidin were mixed thoroughly to obtain a mycotoxin detoxifier for improving mitochondrial function (MDIM). The addition amount in piglet feed was 0.045 wt.%. For ease of subsequent description, this experimental group is designated as the MDIM group.

[0059] Two hundred and forty Duroc-Landrace-Large White piglets with an average weight of 12.3 kg were randomly divided into ten groups: ① Control group (basal diet contaminated with mycotoxins); ② L-malic acid group; ③ α-ketoglutarate group; ④ itaconic acid group; ⑤ 5-norhesperidin group; ⑥ α-ketoglutarate + itaconic acid + 5-norhesperidin group; ⑦ L-malic acid + itaconic acid + 5-norhesperidin group; ⑧ L-malic acid + α-ketoglutarate + 5-norhesperidin group; ⑨ L-malic acid + α-ketoglutarate + itaconic acid group; ⑩ MDIM group. The specific procedures for each group are as follows:

[0060] ① Control group: Feed contaminated with mycotoxins, the contents of aflatoxin B1, vomitoxin, zearalenone and fumonisin in the feed were 6, 845, 141 and 3675 μg / kg, respectively;

[0061] ②L-malic acid group: Based on the control group, L-malic acid was added alone to the same mycotoxin-contaminated feed;

[0062] ③ α-Ketoglutarate group: In addition to the control group, α-ketoglutarate was added alone to the same mycotoxin-contaminated feed;

[0063] ④ Itaconic acid group: In addition to the control group, itaconic acid was added alone to the same mycotoxin-contaminated feed;

[0064] ⑤ The 5-norhesperidin group was supplemented with 5-norhesperidin alone in the same mycotoxin-contaminated feed, in addition to the control group.

[0065] ⑥ α-Ketoglutarate + Itaconic acid + 5-norhesperidin group: Based on the control group, α-ketoglutarate, itaconic acid and 5-norhesperidin were added to the same mycotoxin-contaminated feed.

[0066] ⑦ L-malic acid + itaconic acid + 5-norhesperidin group: Based on the control group, L-malic acid, itaconic acid and 5-norhesperidin were added to the same mycotoxin-contaminated feed.

[0067] ⑧L-malic acid + α-ketoglutarate + 5-norhesperidin group: Based on the control group, L-malic acid, α-ketoglutarate and 5-norhesperidin were added to the same mycotoxin-contaminated feed.

[0068] ⑨ L-malic acid + α-ketoglutarate + itaconic acid group: Based on the control group, L-malic acid, α-ketoglutarate and itaconic acid were added to the same mycotoxin-contaminated feed.

[0069] ⑩ MDIM group: This refers to the addition of the aforementioned composition MDIM in this embodiment, which contains all four components, with an MDIM addition amount of 0.045 wt.% in the feed.

[0070] Except for the control group and the MDIM group, the content of each component added to groups ② to ⑨ above was consistent with the content of the corresponding component in the feed of the MDIM group. For the above ten groups, there were 3 replicates per group, with 8 piglets per replicate. The experimental period was 35 days, and the average daily weight gain and feed conversion ratio of the pigs were calculated. After the feeding trial, 6 pigs from each group were slaughtered, and intestinal, liver, and blood samples were collected for testing. The results of each indicator are as follows: Figure 4 and Figure 5 As shown.

[0071] Depend on Figure 4 As can be seen, compared with the control group, there were no significant differences in daily weight gain, feed conversion ratio, diarrhea rate, villus height-crypt depth ratio (villus-crypt ratio), serum diamine oxidase (DAO), serum aspartate aminotransferase (AST) and alanine aminotransferase (ALT), serum total antioxidant capacity (TAC) and malondialdehyde (MDA), and serum TNF-α in the single-drug groups ②-⑤ and any three-component combination groups ⑥-⑨ (P>0.05). This indicates that single or three-component combinations cannot effectively alleviate mycotoxin-induced growth inhibition, intestinal and liver damage, oxidative stress, and inflammation. However, when the four components were used in combination (MDIM group), all of the above indicators were significantly improved (P<0.05). Bliss independence model analysis showed that the four components had a very strong synergistic effect, which far exceeded the simple sum of the effects of the individual drugs. In summary, L-malic acid, α-ketoglutarate, itaconic acid, and 5-norhesperidin, when combined in specific ratios, exhibit a significant synergistic effect in alleviating mycotoxin-induced growth inhibition, intestinal damage, liver function impairment, oxidative stress, and inflammation, producing unexpected technical benefits.

[0072] Depend on Figure 5 As can be seen, compared with the control group, there were no significant differences in mitochondrial membrane potential (ΔΨm), mitochondrial ROS, respiratory chain complex I activity, cellular oxygen consumption rate (OCR), basal respiration (BR), maximal respiration (MR), ATP production, expression of the glycolysis-related gene PFKFB3, extracellular acidification rate (ECAR), and intestinal mucosal lactate content in each single-drug group ②~⑤ and any three-component combination group ⑥~⑨. Furthermore, morphological changes in intestinal mucosal and liver mitochondria (such as swelling, vacuolation, and cristae damage) were observed in all treatment groups (①-⑨). This indicates that neither a single component nor a combination of three components can effectively alleviate mitochondrial energy metabolism disorders and functional damage caused by mycotoxins. However, when the four components were used in combination (⑩MDIM group), all the above indicators were significantly improved (P<0.05), and no morphological changes in mitochondrial damage were observed in the intestinal mucosa and liver. Bliss independence model analysis showed that the four components had a very strong synergistic effect, far exceeding the simple summation of the effects of each single drug. In summary, L-malic acid, α-ketoglutarate, itaconic acid, and 5-norhesperidin, when combined in specific ratios, exhibit significant synergistic effects on mycotoxin-induced mitochondrial energy metabolism disorders and functional impairment. They can significantly enhance mitochondrial oxidative phosphorylation, reduce glycolysis, improve mitochondrial energy metabolism, and alleviate intestinal and liver mitochondrial damage, producing unexpected technical effects.

[0073] Example 3

[0074] In this embodiment, 13 kg of L-malic acid, 4 kg of calcium α-ketoglutarate, 15 kg of itaconic acid, and 3 kg of 5-norhesperidin were mixed thoroughly to obtain a mycotoxin detoxifier for improving mitochondrial function (MDIM). The addition amount in broiler feed was 0.04 wt.%. For ease of subsequent description, this experimental group is referred to as the MDIM group.

[0075] Four hundred and fifty one-day-old yellow-feathered broilers were randomly divided into ten groups: ① Control group (basal diet contaminated with mycotoxins); ② L-malic acid group; ③ α-ketoglutarate calcium group; ④ itaconic acid group; ⑤ 5-norhesperidin group; ⑥ α-ketoglutarate calcium + itaconic acid + 5-norhesperidin group; ⑦ L-malic acid + itaconic acid + 5-norhesperidin group; ⑧ L-malic acid + α-ketoglutarate calcium + 5-norhesperidin group; ⑨ L-malic acid + α-ketoglutarate calcium + itaconic acid group; ⑩ MDIM group. The specific procedures for each group are as follows:

[0076] ① Control group: Feed contaminated with mycotoxins, the contents of aflatoxin B1, vomitoxin, zearalenone and T-2 toxin in the feed were 8, 1608, 406 and 450 μg / kg, respectively;

[0077] ②L-malic acid group: Based on the control group, L-malic acid was added alone to the same mycotoxin-contaminated feed;

[0078] ③Calcium α-ketoglutarate group: In addition to the control group, calcium α-ketoglutarate was added alone to the same mycotoxin-contaminated feed;

[0079] ④ Itaconic acid group: In addition to the control group, itaconic acid was added alone to the same mycotoxin-contaminated feed;

[0080] ⑤ The 5-norhesperidin group was supplemented with 5-norhesperidin alone in the same mycotoxin-contaminated feed, in addition to the control group.

[0081] ⑥ α-Ketoglutarate calcium + itaconic acid + 5-norhesperidin group: Based on the control group, α-ketoglutarate calcium, itaconic acid and 5-norhesperidin were added to the same mycotoxin-contaminated feed.

[0082] ⑦ L-malic acid + itaconic acid + 5-norhesperidin group: Based on the control group, L-malic acid, itaconic acid and 5-norhesperidin were added to the same mycotoxin-contaminated feed.

[0083] ⑧L-malic acid + α-ketoglutarate calcium + 5-norhesperidin group: Based on the control group, L-malic acid, α-ketoglutarate calcium, and 5-norhesperidin were added to the same mycotoxin-contaminated feed.

[0084] ⑨ L-malic acid + α-ketoglutarate calcium + itaconic acid group: Based on the control group, L-malic acid, α-ketoglutarate calcium, and itaconic acid were added to the same mycotoxin-contaminated feed.

[0085] ⑩ MDIM Group: This group included the aforementioned composition MDIM from this embodiment, containing all four components, added to the feed at a rate of 0.04 wt.%. The content of the added components in the feed of each group was consistent with the content of the corresponding components in the feed of the MDIM group. Each group had three replicates, with 15 birds per replicate, and the trial lasted 70 days. The average daily weight gain and feed conversion ratio of the broilers were calculated. After the feeding trial, six broilers from each group were slaughtered, and intestinal, liver, and blood samples were collected for testing. The results of each indicator are as follows: Figure 6 and Figure 7 As shown.

[0086] Depend on Figure 6 It is evident that there were no significant differences in daily weight gain and feed conversion ratio between the single-drug groups and any three-component combination groups and the control group (P>0.05), indicating that neither single nor three-component combinations could effectively alleviate growth inhibition caused by mycotoxins. However, when all four components were used in combination (MDIM group), daily weight gain was significantly increased and feed conversion ratio was significantly decreased (P<0.05). Bliss independence model analysis showed that the four components had a very strong synergistic effect, far exceeding the simple sum of the effects of the individual drugs. In conclusion, L-malic acid, calcium α-ketoglutarate, itaconic acid, and 5-norhesperidin, in specific ratios, have a significant synergistic effect on alleviating growth inhibition caused by mycotoxins, producing unexpected technical effects on both daily weight gain and feed conversion ratio, two independent indicators.

[0087] Depend on Figure 6It is evident that, compared with the control group, there were no significant differences in the chorionic villus ratio, serum diamine oxidase (DAO), serum aspartate aminotransferase (AST) and alanine aminotransferase (ALT), serum total antioxidant capacity (TAC) and malondialdehyde (MDA), and serum TNF-α in any single-drug group or any combination of three components (P>0.05). This indicates that single or three-component combinations cannot effectively alleviate mycotoxin-induced intestinal and liver damage, oxidative stress, and inflammation. However, when all four components were used in combination (MDIM group), all of the above indicators were significantly improved (P<0.05). Bliss independence model analysis showed that the four components had a very strong synergistic effect, far exceeding the simple sum of the effects of the individual drugs. In conclusion, L-malic acid, calcium α-ketoglutarate, itaconic acid, and 5-norhesperidin, in a specific ratio, have a significant synergistic effect on alleviating mycotoxin-induced intestinal and liver damage, oxidative stress, and inflammation, producing unexpected technical effects.

[0088] Depend on Figure 7 As can be seen, compared with the control group, there were no significant differences in mitochondrial membrane potential (ΔΨm), mitochondrial ROS, respiratory chain complex I activity, cellular oxygen consumption rate (OCR), basal respiration (BR), maximal respiration (MR), ATP production, glycolysis-related gene PFKFB3 expression, extracellular acidification rate (ECAR), and intestinal mucosal lactate content in each single-drug group and any combination of three components (P>0.05). Furthermore, morphological changes in intestinal mucosal and liver mitochondria (such as swelling, vacuolation, and cristae damage) were observed in all treatment groups (①-⑨). This indicates that neither a single component nor a combination of three components can effectively alleviate mitochondrial energy metabolism disorders and functional damage caused by mycotoxins. However, when the four components were used in combination (⑩MDIM group), all the above indicators were significantly improved (P<0.05), and no morphological changes in mitochondrial damage were observed in the intestinal mucosa and liver. Bliss independence model analysis showed that the four components had a very strong synergistic effect, far exceeding the simple sum of the effects of the individual drugs. In summary, L-malic acid, calcium α-ketoglutarate, itaconic acid, and 5-norhesperidin, when combined in specific ratios, exhibit significant synergistic effects on mycotoxin-induced mitochondrial energy metabolism disorders and functional impairment. They can significantly enhance mitochondrial oxidative phosphorylation, reduce glycolysis, improve mitochondrial energy metabolism, and alleviate intestinal and liver mitochondrial damage, producing unexpected technical effects.

[0089] The embodiments described above are merely some preferred implementations of the present invention and are not intended to limit the invention. Those skilled in the art can make various changes and modifications without departing from the spirit and scope of the invention. Therefore, all technical solutions obtained through equivalent substitution or transformation fall within the protection scope of the present invention.

Claims

1. A mycotoxin attenuator that improves mitochondrial function, characterized in that, It includes the following components in parts by weight: 10-20 parts of L-malic acid, 3-5 parts of α-ketoglutarate or α-ketoglutarate, 15-25 parts of itaconic acid, and 1-3 parts of 5-norhesperidin.

2. The mycotoxin attenuator for improving mitochondrial function according to claim 1, characterized in that, The mycotoxin attenuator, by weight, is composed of 10-20 parts of L-malic acid, 3-5 parts of α-ketoglutarate or α-ketoglutarate salt, 15-25 parts of itaconic acid, and 1-3 parts of 5-norhesperidin.

3. The mycotoxin attenuator for improving mitochondrial function according to claim 1, characterized in that, The α-ketoglutarate is sodium α-ketoglutarate or calcium α-ketoglutarate.

4. A feed containing a mycotoxin attenuator as described in any one of claims 1 to 3 that improves mitochondrial function.

5. The feed as described in claim 4, characterized in that, The amount of mycotoxin attenuator added to the feed is 0.04 wt.%~0.05 wt.%.

6. The feed as described in claim 4, characterized in that, The feed is pig feed or broiler feed.

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