Mixed traditional Chinese medicine feed additive for resisting jejunum, white stool and intestinal erosion of fishes

Abstract

The invention belongs to the technical field of animal feed, and particularly relates to a mixed traditional Chinese medicine feed additive for resisting jejunum, white stool and intestinal erosion of fishes, which is prepared from the following raw material components: sea-buckthorn, isatis tinctoria and safflower. The sea-buckthorn, the isatis tinctoria and the safflower are mixed and crushed, and then water extraction and probiotic fermentation treatment are performed, so that the obtained mixed traditional Chinese medicine feed additive can effectively improve the symptoms of jejunum, white stool and intestinal erosion of fishes, and is a natural traditional Chinese medicine feed additive which is safe, effective and free of toxic and side effects; and the urgent demand in the current aquaculture field is met.

Description

Technical Field

[0001] The invention belongs to the technical field of animal feed, and in particular relates to a mixed Chinese medicinal feed additive for resisting jejunal, white stool and intestinal erosion of fish. Background Art

[0002] The digestive system of fish is easily affected by various factors, leading to jejunal indigestion, white stool, intestinal erosion and other diseases. Especially in the case of unstable water quality or poor feed quality, the incidence of these diseases is higher.

[0003] Traditional prevention and treatment methods mostly rely on antibiotics and chemical drugs, but long-term use may lead to drug residues, environmental pollution and drug resistance. Chinese herbal medicines, as natural medicines, have a long history in the prevention and treatment of aquatic animal diseases. Because of their advantages such as small toxic side effects, less residues, and no obvious drug resistance, in addition to their medicinal properties, they can also play a preventive and therapeutic role through their nutritional effects, immune enhancement effects, and hormone-like effects on aquatic animals. Therefore, the development of a safe, effective, and non-toxic natural Chinese herbal feed additive has become an urgent need in the current aquaculture field. Summary of the invention

[0004] In order to solve the above problems, in the first aspect, the present invention provides a mixed Chinese medicinal feed additive for preventing jejunal, white stool and intestinal erosion in fish, wherein the mixed Chinese medicinal feed additive is prepared from the following raw material components: sea buckthorn, isatis indigotica and safflower.

[0005] Furthermore, the weight proportions of the raw material components are: 0.5-5 parts of seabuckthorn; 0.5-5 parts of isatis indigotica; and 0.5-5 parts of safflower.

[0006] Furthermore, the weight proportions of the raw material components are: 1 part of seabuckthorn; 1 part of isatis indigotica; and 1 part of safflower.

[0007] In a second aspect, the present invention provides a method for preparing a mixed Chinese medicinal feed additive for preventing fish jejunum, white stool and intestinal erosion as described herein, comprising the following steps:

[0008] 1) Weigh seabuckthorn, isatis indigotica and safflower according to the required weight, mix and grind to obtain a mixed powder;

[0009] 2) extracting the mixed powder with water to obtain a water extract;

[0010] 3) Mixing the obtained water extract with fermentation medium, inoculating fermentation bacteria for fermentation, and obtaining the product.

[0011] Furthermore, the water extraction comprises adding the mixed powder into 8-12 times the weight of water, heating and boiling for extraction, filtering, and taking the supernatant to obtain a water extract.

[0012] Furthermore, the fermentation bacteria are Bacillus subtilis and yeast.

[0013] Furthermore, the inoculation amount of the Bacillus subtilis and the yeast is 1-5wt%;

[0014] Furthermore, the accession amount of the Bacillus subtilis is 2wt%, and the accession amount of the yeast is 5wt%.

[0015] Furthermore, the fermentation includes continuous stirring for 12-48 hours at 28-30° C., and during the stirring process, air is introduced into the fermentation tank to perform aerobic fermentation.

[0016] Furthermore, the fermentation medium and the water extract are mixed in a volume ratio of 1:1.

[0017] Furthermore, the formula of the fermentation medium is: 10g of refined peptone, 40g of glucose, 5g of sodium acetate, 2g of ammonium citrate, 0.1g of Tween 80, 0.58g of magnesium sulfate, 0.28g of manganese sulfate, and 1000mL of distilled water; the pH is adjusted to 6-7 with 10% sodium hydroxide solution, and high-pressure sterilization is performed at 121°C for 20min.

[0018] Furthermore, the mixed Chinese medicinal feed additive for preventing fish jejunal, white stool and intestinal erosion prepared by the method can be used as a fluid dosage form, or can be dried, such as freeze-dried, to form a powder, preferably freeze-dried to form a powder.

[0019] Furthermore, when used, the mixed Chinese medicinal feed additive provided by the present invention is added to the basic feed of fish. Furthermore, the mixed Chinese medicinal feed additive provided by the present invention is added to the basic feed of fish at an addition ratio of 1-10wt%.

[0020] Therefore, in another aspect, the present invention provides a fish feed, which comprises the mixed Chinese medicinal feed additive for resisting fish jejunum, white stool and intestinal erosion according to the present invention; the addition amount thereof is 1-10wt%.

[0021] Beneficial effects of the invention:

[0022] (1) Seabuckthorn, Isatis indigotica and Safflower are commonly used anti-inflammatory Chinese medicines, but their use in fish feed has not been reported. The present invention has found that the combined use of seabuckthorn, Isatis indigotica and Safflower can effectively improve the symptoms of jejunum, white stool and intestinal erosion in fish.

[0023] (2) In order to improve the efficacy of the medicine, the present invention extracts the Chinese medicine formula with water, and then ferments the Chinese medicine with probiotics. The macromolecular substances in the Chinese medicine can be transformed into small molecular components that can be directly absorbed by the body's intestines through microorganisms, which can increase the precipitation of effective ingredients of the Chinese medicine such as ketones, esters and other substances, improve the drug absorption capacity, help improve the biotransformation effect and the utilization rate of medicinal materials, and significantly enhance the improvement effect on the fish digestive system, without causing fish jejunal, white stool, intestinal erosion and other diseases. DETAILED DESCRIPTION

[0024] The present invention is further described below with reference to specific examples, but the examples do not limit the present invention in any form. Unless otherwise specified, the reagents, methods and equipment used in the present invention are conventional reagents, methods and equipment in the art.

[0025] In the following examples, yeast was purchased from China Industrial Microorganism Culture Collection Administration Center, strain number: CICC 32883. Bacillus subtilis was purchased from China Industrial Microorganism Culture Collection Administration Center, strain number: CICC25064.

[0026] Before the experiment, the purchased yeast and Bacillus subtilis freeze-dried powder were inoculated into the fermentation medium respectively and cultured in a constant temperature shaking incubator at a speed of 200 r / min and 30°C for 24 h.

[0027] The formula of the fermentation medium is: 10g of refined peptone, 40g of glucose, 5g of sodium acetate, 2g of ammonium citrate, 0.1g of Tween 80, 0.58g of magnesium sulfate, 0.28g of manganese sulfate, and 1000mL of distilled water; the pH is adjusted to 6-7 with 10% sodium hydroxide solution, and the medium is sterilized by high pressure at 121°C for 20min.

[0028] Embodiment 1:

[0029] This embodiment provides a mixed Chinese medicinal feed additive for preventing fish jejunal, white stool, and intestinal erosion, which is prepared as follows:

[0030] 1) Weigh 15 g of seabuckthorn, 15 g of isatis indigotica and 15 g of safflower, mix and grind to obtain a mixed powder;

[0031] 2) Add the mixed powder to an extraction tank, add 10 times the weight of water, heat and boil for 1 hour, and filter; continue to extract the filter residue for 2 times; combine the extracts from the 3 times to obtain the water extracts of seabuckthorn, isatis indigotica and safflower;

[0032] 3) The obtained water extract was mixed with the fermentation medium in a volume ratio of 1:1, and Bacillus subtilis (inoculation amount of 2 wt%) and yeast (inoculation amount of 5 wt%) were inoculated, and the mixture was stirred at 250 r / min and 0.05 m 3 / min ventilation rate, fermentation temperature of 30 ℃ for 48h, and freeze-drying to obtain the product.

[0033] Embodiment 2:

[0034] This embodiment provides a mixed Chinese medicinal feed additive for preventing fish jejunal, white stool, and intestinal erosion, which is prepared as follows:

[0035] 1) Weigh 20 g of seabuckthorn, 10 g of isatis indigotica and 30 g of safflower, mix and grind to obtain a mixed powder;

[0036] 2) Add the mixed powder to an extraction tank, add 10 times the weight of water, heat and boil for 1 hour, and filter; continue to extract the filter residue for 2 times; combine the extracts from the 3 times to obtain the water extracts of seabuckthorn, isatis indigotica and safflower;

[0037] 3) The obtained water extract was mixed with the fermentation medium in a volume ratio of 1:1, and Bacillus subtilis (inoculation amount of 1wt%) and yeast (inoculation amount of 5wt%) were inoculated, and the mixture was stirred at 250r / min and 0.05m 3 / min ventilation rate, fermentation temperature of 30 ℃ for 48h, and freeze-drying to obtain the product.

[0038] Comparative Example 1:

[0039] This comparative example provides a Chinese medicinal feed additive, which is prepared as follows:

[0040] 1) Weigh 45 g of sea buckthorn and crush it to obtain powder;

[0041] 2) Same as Example 1;

[0042] 3) Same as Example 1.

[0043] Comparative Example 2:

[0044] This comparative example provides a Chinese medicinal feed additive, which is prepared as follows:

[0045] 1) Weigh 45 g of Isatis indigotica and crush it to obtain a powder;

[0046] 2) Same as Example 1;

[0047] 3) Same as Example 1.

[0048] Comparative Example 3:

[0049] This comparative example provides a Chinese medicinal feed additive, which is prepared as follows:

[0050] 1) Weigh 45 g of safflower and crush it to obtain powder;

[0051] 2) Same as Example 1;

[0052] 3) Same as Example 1.

[0053] Comparative Example 4:

[0054] This comparative example provides a mixed Chinese medicinal feed additive, which is prepared as follows:

[0055] 1) Weigh 20 g of seabuckthorn and 20 g of safflower, mix and grind to obtain a mixed powder;

[0056] 2) Same as Example 1;

[0057] 3) Same as Example 1.

[0058] Comparative Example 5:

[0059] This comparative example provides a Chinese medicinal feed additive, which is prepared as follows:

[0060] 1) Weigh 45 g of sea buckthorn and crush it to obtain powder;

[0061] 2) Add the powder to an extraction tank, add 10 times the weight of water, heat and boil for 1 hour, and filter; continue to extract the filter residue for 2 more times; combine the extracts from the 3 times to obtain the water extracts of seabuckthorn, isatis indigotica and safflower, and freeze-dry.

[0062] Comparative Example 6:

[0063] This comparative example provides a Chinese medicinal feed additive, which is prepared as follows:

[0064] 1) Weigh 45 g of Isatis indigotica and crush it to obtain a powder;

[0065] 2) Add the powder to an extraction tank, add 10 times the weight of water, heat and boil for 1 hour, and filter; continue to extract the filter residue for 2 more times; combine the extracts from the 3 times to obtain the water extracts of seabuckthorn, isatis indigotica and safflower, and freeze-dry.

[0066] Comparative Example 7:

[0067] This comparative example provides a Chinese medicinal feed additive, which is prepared as follows:

[0068] 1) Weigh 45 g of safflower and crush it to obtain powder;

[0069] 2) Add the powder to an extraction tank, add 10 times the weight of water, heat and boil for 1 hour, and filter; continue to extract the filter residue for 2 more times; combine the extracts from the 3 times to obtain the water extracts of seabuckthorn, isatis indigotica and safflower, and freeze-dry.

[0070] Comparative Example 8:

[0071] This comparative example provides a Chinese medicinal feed additive, which is prepared as follows:

[0072] 1) Weigh 20 g of seabuckthorn and 20 g of safflower, mix and grind to obtain a mixed powder;

[0073] 2) Add the mixed powder into an extraction tank, add 10 times the weight of water, heat and boil for 1 hour, and filter; continue to extract the filter residue for 2 more times; combine the extracts from the 3 times to obtain the water extracts of seabuckthorn, isatis indigotica and safflower, and freeze-dry.

[0074] Comparative Example 9:

[0075] This comparative example provides a mixed Chinese medicinal feed additive, which is prepared as follows:

[0076] 1) Weigh 15 g of seabuckthorn, 15 g of isatis indigotica and 15 g of safflower, mix and grind to obtain a mixed powder;

[0077] 2) Add the mixed powder into an extraction tank, add 10 times the weight of water, heat and boil for 1 hour, and filter; continue to extract the filter residue for 2 more times; combine the extracts from the 3 times to obtain the water extracts of seabuckthorn, isatis indigotica and safflower, and freeze-dry.

[0078] Test example:

[0079] Healthy, undamaged, uniform-sized turbot seedlings weighing 3-5 g were selected at the breeding base and fed with a basic feed (special feed for turbot, purchased from Shandong Hanye Biotechnology Co., Ltd.) supplemented with 1 wt% of the additives of Examples 1-2 or Comparative Examples 1-9 for 2 weeks. After the feeding of the feed containing the additive was completed, the basic feed was continued for 8 weeks.

[0080] After feeding, Edwardsiella piscicola EIB202 was used to challenge turbot. In brief, EIB202 bacterial solution was inoculated into TYB medium containing 50 μg / ml polymyxin at a 1% inoculation rate and cultured at 28°C and 200 rpm. The bacterial concentration was measured by NanoDrop2000, and when OD600 increased to 4, the bacteria were collected by centrifugation at 8000 rpm for 10 min. 40 turbot were randomly selected from each group for soaking and challenge, the soaking volume was 6 L, and the bacterial concentration was 2×10 8 CFU / ml, immersion time was 1h. After immersion, it was washed with seawater and placed in a culture tank for subsequent observation.

[0081] Seven days after bacterial infection, the pathological phenotypes of jejunum, white feces and intestinal erosion in the intestinal tract of turbot seedlings in different feeding groups were observed. The results are shown in Table 1.

[0082] Table 1: Incidence of jejunum, white stool and intestinal erosion in turbot

[0083]

[0084]

[0085] From the results in Table 1, it can be seen that after the infection, the turbot in the blank control group showed more symptoms of jejunum, white stool and intestinal erosion, but after being fed with the feed added with the example, the symptoms of jejunum, white stool and intestinal erosion were significantly improved.

[0086] From the results of Comparative Examples 1-4, it can be seen that when the additive is prepared from only one of seabuckthorn, woad and safflower, the pathological condition of turbot is slightly alleviated, among which seabuckthorn and safflower have better effects. When these two are combined (Comparative Example 4), it can be seen that the relief of the pathological condition of turbot is significantly improved, indicating that seabuckthorn and safflower can synergistically improve the symptoms of jejunum, white stool and intestinal erosion of turbot. On this basis, adding woad extract (Example 1) has a more significant improvement effect.

[0087] From the results of Comparative Examples 5-9, it can be seen that when preparing additives, only water extraction is used to extract Chinese medicine, and probiotic fermentation is not used to assist. Whether it is a single Chinese medicine raw material or a combination of the three, the improvement effect on the symptoms of turbot jejunum, white stool and intestinal erosion is not as good as the effect of probiotic fermentation. An interesting discovery is that when only the water extraction process is used, the combination of seabuckthorn and safflower can still see obvious synergistic effects (Comparative Example 8), but on this basis, the improvement effect of the water extract of the combination of the three with the addition of Isatis indigotica is not significantly improved (Comparative Example 9), while in contrast, when the fermentation process is also used, the addition of Isatis indigotica significantly improves the effect of the combination of seabuckthorn and safflower, indicating that the fermentation treatment of Isatis indigotica is indispensable for the efficacy of the feed additive of the present invention.

[0088] It should be noted that the preferred embodiments of the present invention are given in the specification of the present invention, but the present invention can be implemented in many different forms and is not limited to the embodiments described in this specification. These embodiments are not intended to be additional limitations on the content of the present invention. The purpose of providing these embodiments is to make the understanding of the disclosure of the present invention more thorough and comprehensive. In addition, the above-mentioned technical features continue to be combined with each other to form various embodiments not listed above, which are all considered to be within the scope of the present invention specification; further, for ordinary technicians in this field, they can be improved or transformed according to the above description, and all these improvements and transformations should belong to the protection scope of the claims attached to the present invention.

Claims

1. A mixed Chinese medicinal feed additive for preventing fish jejunum, white stool and intestinal erosion, characterized in that: The mixed Chinese medicinal feed additive is prepared from the following raw material components: seabuckthorn, isatis indigotica and safflower.

2. The mixed Chinese medicinal feed additive according to claim 1, characterized in that: The weight proportions of the raw material components are: 0.5-5 parts of seabuckthorn, 0.5-5 parts of isatis indigotica, and 0.5-5 parts of safflower.

3. The mixed Chinese medicinal feed additive according to claim 1, characterized in that: The weight proportions of the raw material components are: 1 part of seabuckthorn; 1 part of isatis indigotica; and 1 part of safflower.

4. A method for preparing a mixed Chinese medicinal feed additive for preventing jejunal, fecal ulcer and intestinal erosion in fish as claimed in any one of claims 1 to 3, characterized in that: The following steps are involved: 1) Weigh seabuckthorn, isatis indigotica and safflower according to the required weight, mix and grind to obtain a mixed powder; 2) extracting the mixed powder with water to obtain a water extract; 3) Mixing the obtained water extract with fermentation medium, inoculating fermentation bacteria for fermentation, and obtaining the product.

5. The method according to claim 4, characterized in that The water extraction comprises adding the mixed powder into 8-12 times the weight of water, heating and boiling for extraction, filtering, and taking the supernatant to obtain a water extract.

6. The method according to claim 4, characterized in that The fermentation bacteria are Bacillus subtilis and yeast; The inoculation amount of the Bacillus subtilis and yeast is 1-5wt%; The inoculation amount of the Bacillus subtilis is 2 wt %, and the inoculation amount of the yeast is 5 wt %.

7. The method according to claim 4, characterized in that The fermentation includes continuous stirring for 12-48 hours at 28-30° C., during which air is introduced into the fermentation tank for aerobic fermentation; After the fermentation is completed, the fermentation liquid is freeze-dried to obtain a mixed Chinese medicine feed additive powder.

8. The method according to claim 4, characterized in that The fermentation medium and the water extract are mixed in a volume ratio of 1:

1.

9. The method according to claim 4, characterized in that The formula of the fermentation medium is: 10g of refined peptone, 40g of glucose, 5g of sodium acetate, 2g of ammonium citrate, 0.1g of Tween 80, 0.58g of magnesium sulfate, 0.28g of manganese sulfate, and 1000mL of distilled water; the pH is adjusted to 6-7 with 10% sodium hydroxide solution, and the medium is sterilized by high pressure at 121°C for 20min.

10. A fish feed, characterized in that: A mixed Chinese medicinal feed additive for resisting jejunum, white stool and intestinal erosion of fish according to any one of claims 1 to 3 or a mixed Chinese medicinal feed additive for resisting jejunum, white stool and intestinal erosion of fish prepared by the method according to any one of claims 4 to 9.

Images