CN115487206A

CN115487206A - Method for relieving poor development of tibial cartilage of broiler chicken by morinda officinalis polysaccharide and application of morinda officinalis polysaccharide

Info

Publication number: CN115487206A
Application number: CN202211245466.9A
Authority: CN
Inventors: 黄淑成; 张朝栋; 郭娜; 刘芳; 刘凯莉
Original assignee: Henan Agricultural University
Current assignee: Henan Agricultural University
Priority date: 2022-10-08
Filing date: 2022-10-08
Publication date: 2022-12-20

Abstract

The morinda officinalis polysaccharide can relieve lameness of the broiler caused by the tibial cartilage dysplasia, improves related indexes of the tibia through a BMP/Smads signal path, restores the structure of a tibial growth plate and balances calcium and phosphorus metabolic disorder caused by TD. Meanwhile, the morinda officinalis polysaccharide also improves the oxidation resistance of organisms, improves the structure of intestinal flora of the broiler chickens, improves the abundance of beneficial bacteria and reduces the abundance of harmful bacteria.

Description

Method for relieving poor development of tibial cartilage of broiler chicken by morinda officinalis polysaccharide and application of morinda officinalis polysaccharide

Technical Field

The invention relates to a novel method and application of morinda officinalis polysaccharide, in particular to a method for relieving poor development of tibial cartilage of a broiler and application of morinda officinalis polysaccharide.

Background

Tibial Dyschondroplasia (TD) is a common nutritional metabolic leg disease in rapidly-forming poultry. The disease is characterized by avascular and jade-white cartilage wedges formed by the ossification blockage in cartilage and the proliferation of chondrocytes at the metaphysis of tibia, and has the clinical manifestations of listlessness, appetite reduction, unwilling standing, difficult walking, tibial development deformity and the like, which cause the reduction of the production performance and the low immune function of broilers, and can also induce a series of secondary diseases such as chest cyst, fracture, osteomyelitis and the like to influence the body health of the broilers. The disease is firstly discovered and reported in 1965 in a fast-growing broiler chicken by Leach RM and MC Nesheim in the United states, and then is discovered in succession in the breeding process of poultry such as ducks, turkeys and the like. With the development of global economy, the demand of poultry meat and egg products is increased, and in the process of pursuing development quantity, yield and marketing speed, the leg diseases of the broilers are increased due to the limitation of feeding space and the over-high weight increasing speed in an intensive chicken raising mode, so that the serious threat to the health of the broilers is formed, and the global loss is brought to the broiler breeding industry.

At present, no specific medicine which can be widely used for treating TD exists. Previous studies show that the incidence of TD in broiler chickens can be reduced by feeding vitamin C and vitamin D3 and changing the ratio of calcium to phosphorus in the diet. However, because the production period of broiler chickens is short, the metabolism period of medicines is long, the treatment cost is high, and medicines are remained in commercial chickens and are accumulated continuously, how to widely use the medicines to prevent and treat TD is an important problem.

The traditional Chinese medicine has a long application history, and is widely used for preventing and treating diseases due to good curative effect, small toxicity and small side effect. Morinda citrifolia polysaccharide (MOP) is a derivative extracted from Morinda citrifolia, widely distributed in nature, having antioxidant, anti-inflammatory and bone formation promoting effects. Research shows that MOP can effectively improve the bone density of rats with ovariectomy osteoporosis, and has the effect of resisting osteoporosis by regulating the levels of Tumor necrosis factor-alpha (TNF-alpha) and Interleukin-1 (Interleukin-1, IL-1) in the rats. Meanwhile, MOP also has a certain effect on improving the mineral content in ovariectomized rats, and the MOP can improve the contents of bone calcium and bone phosphorus by regulating the levels of nuclear factor-kappa B Receptor activating factors (RANK) and Osteoprotegerin (OPG), so that the effect of reducing the generation of osteoclasts is achieved. MOP may also promote bone formation by modulating the BMP-2/Runx-2/Osterix pathway. In view of the effect of MOP on the bone growth and development of animals, MOP may have certain potential in preventing and treating bone growth and development diseases of broilers.

Disclosure of Invention

Experiments show that MOP can relieve lameness of broiler chickens caused by TD, improve relevant indexes of shin bones through a BMP/Smads signal path, recover the structure of the shin bone growth plate and balance calcium and phosphorus metabolism disorder caused by TD. Meanwhile, the MOP also improves the oxidation resistance of organisms, improves the structure of intestinal flora of the broiler chickens, improves the abundance of beneficial bacteria, reduces the abundance of harmful bacteria, provides a new idea for treating TD, and proves the potential application value of the MOP in preventing and treating TD.

The invention adopts Yang Lingci and morinda officinalis polysaccharide provided by biotechnology limited company, and the morinda officinalis polysaccharide which is a traditional Chinese medicine extract is used for broiler chickens. Preferably, the morinda officinalis how-glucose is administered in an amount of 500mg/kg in drinking water.

The invention utilizes Ai Ba Yijia (Arbor acids, AA) broiler chicken to establish a thiram induced TD model, the TD model is induced by feeding 100mg/kg thiram added feed in the 4 th to 7 th days, and the incidence rate is obviously higher than that caused by daily ration component change, acid-base balance change and other factors. And the TD model induced by thiram can ensure that the broiler chickens are attacked in the same period on one hand and the incidence rate reaches 100 percent on the other hand, thereby providing sufficient and reliable test materials for deeply researching the pathogenesis of the broiler chickens and being used as an ideal test animal model for researching the broiler chickens. The morinda officinalis how polysaccharide is administered in water at a dose of 500mg/kg/d for fourteen consecutive days. Body weight was recorded daily and broiler clinical symptoms were observed. The method comprises the following steps of taking materials after 14 days, recording the weight and length of the tibia and the width of a tibia growth plate, measuring the ash content, calcium content and phosphorus content in the tibia, extracting broiler tibia growth plate RNA, carrying out RT-qPCR to measure the expression level of BMP/Smads related genes, carrying out histopathological examination on the tibia growth plate, measuring the levels of calcium, phosphorus and bone formation marker alkaline phosphatase (ALP) in plasma by adopting blood, and proving that MOP improves the tibia related indexes through a BMP/Smads signal path, restores the tibia growth plate structure and balances the calcium-phosphorus metabolic disorder caused by TD. Through determining the relevant indexes of plasma oxidative stress, the MOP can improve the oxidation resistance of an organism, collects the caeca content of broiler chickens, performs 16s RNA sequencing, performs OUT statistical analysis, alpha diversity analysis, beta diversity analysis and dominant flora analysis on sample intestinal flora, finds that the MOP can be used for TD broiler intestinal flora structure, improves the relevant abundance of beneficial bacteria, reduces the relevant abundance of harmful bacteria, and further regulates and controls the metabolic function of the organism.

Drawings

FIG. 1. Influence of MOP on clinical symptoms and histopathology of TD broiler chickens

(A) The clinical characteristics of the broiler adaptation period, the thiram induction TD period and the MOP treatment period. (B) Tibial and tibial growth plate morphology examination (C) tibial growth plate index and tibial weight index. (D) examination of tibial growth plate histopathology.

FIG. 2 shows the influence of MOP on the ash content, calcium content and phosphorus content of tibia of TD broiler chicken, the calcium and phosphorus level of blood plasma and bone metabolism index.

(A) Calcium, phosphorus and ash contents of tibia on 14 and 21 days. (B) plasma calcium levels. (C) plasma phosphorus levels. (D) plasma ALP levels.

FIG. 3. Influence of MOP on TD broiler BMP/Smads signal pathway

FIG. 4. Influence of MOP on antioxidant capacity of TD broiler chicken

(A) Plasma T-SOD, GSH-Px activity and MDA level. (B) Tibial growth plate SOD, GPX-1 gene expression level

FIG. 5. Influence of MOP on TD broiler intestinal flora

(A) Venn diagram based on OTUs. (B) Beta diversity: PCoA based on Unweighted-UniFrac. (C) Alpha diversity: simpson index, chao1 index, observe _ speces index, shannon index. (D) phylum level intestinal flora composition and relative abundance. (E) genus level intestinal flora composition and relative abundance.

Advantageous effects

1. The invention provides a traditional Chinese medicine extract, namely morinda officinalis polysaccharide, wherein the morinda officinalis polysaccharide is an extract from a traditional Chinese medicine morinda officinalis, is widely distributed in nature, has the effects of resisting oxidation and inflammation and promoting bone formation, and has good curative effect, small toxicity, small side effect and potential clinical treatment value.

2. The in vivo verification proves that the morinda officinalis how polysaccharide has the characteristics of promoting bone growth, resisting oxidation and improving the structure of intestinal flora, can effectively relieve the poor development of tibial cartilage of the broiler chicken, and provides a new idea for preventing and treating TD.

Detailed Description

The specific implementation mode of the invention comprises the following contents:

as a preferable technical scheme, the method comprises a method for improving thiram-induced broiler tibia dysplasia through a BMP/Smads signaling pathway by morinda officinalis polysaccharide and application of the method.

As the preferable technical scheme, the method comprises the steps of improving the antioxidant capacity of TD broiler chickens by morinda officinalis polysaccharides and the application;

as a preferable technical scheme, the method comprises a method for improving the intestinal flora of TD broiler chickens by using morinda officinalis polysaccharides and an application of the morinda officinalis polysaccharides.

Example 1

The method for improving thiram-induced broiler tibia cartilage dysplasia by morinda officinalis polysaccharide through BMP/Smads signal path comprises the following steps:

step one, selection of raw materials and breeding method

Morinda citrifolia polysaccharide (lot number: CY 180812) was purchased from Yang Lingci Yuan Biotechnology, inc. Thiram (tetramethylthiuram disulfide, M03718066) available from shanghai mclin biochemistry ltd. 120 AA chickens (1 day old; 46.96 +/-6.53 g) were selected and obtained from Kyoda poultry industries, inc.

All broilers were housed in standard room temperature and relative humidity light/dark cycle facilities and provided free of water and food. The broiler chickens were randomly divided into 3 groups of 10 chickens each, 4 replicates each, control (CON), thiram-induced TD and MOP treated groups respectively. On the 4 th to 7 th days, the CON group is fed with normal diet, and the TD group and the MOP group are fed with diet added with thiram of 100 mg/kg. On 7-21 days, 500mg/kg of MOP is added into drinking water for the broiler chickens in the MOP treatment group. The test was carried out for a total of 21 days.

Step two, sampling and measuring method

The broiler weight was recorded daily during the test period, 10 chickens per group were sacrificed on

days

7, 14 and 21, respectively, by cervical dislocation, and plasma samples were stored at 4 ℃,3000rpm for 10min and-20 ℃ after carotid vein aseptic blood sampling. Tibia length and growth plate width were measured and H & E stained. The right shin bones of 6 broilers in each group were taken and the ash, calcium and phosphorus contents in the shin bones were measured. Plasma calcium, phosphorus and ALP levels were determined by the kit.

Trizol method for extracting tibial growth plate RNA

Preparing at the early stage of the test: instruments such as a mortar, a grinding rod, a surgical scissors and the like for extracting RNA are sterilized by high-pressure steam in an autoclave in advance, and then are added into liquid nitrogen for precooling.

(1) The tibial growth plate tissue was removed from the-80 ℃ freezer, a small amount (soybean size) of the tissue was placed in a mortar and liquid nitrogen was added, ground with a grinding bar, while liquid nitrogen was continuously added to the mortar until the tissue was ground to a powder, and the sample powder was transferred to a 1.5ml enzyme-free centrifuge tube using a small spatula.

(2) 1mL of Trizol solution (the sample is not sufficiently cracked due to too little lysate, and the concentration of the product is reduced) is added into a 1.5mL EP tube, and the mixture is shaken vigorously by using an oscillator or blown and beaten repeatedly by using a pipette gun, inverted, mixed evenly and placed for 5min.

(3) 200ml of chloroform is added into an EP tube, the mixture is stirred and mixed evenly, and the mixture is placed for 5min at room temperature. C,12000/15min.

(4) The supernatant was aspirated by a pipette gun, isopropanol was added, and the mixture was allowed to stand at room temperature for 10min,4. C12000 and 10min.

(5) The supernatant was discarded and 1ml of 75% ethanol was added for washing, and the mixture was flicked and mixed, 4. C7500 rotating for 5min.

(6) And 6, repeating the step.

(7) Standing at room temperature, air drying, adding 20 μ L of RNase-free ddH2O to dissolve the precipitate, and slightly shaking for 3min to dissolve the RNA precipitate completely.

(8) The concentration of RNA was measured using a nucleic acid protein analyzer and reduced to 1000 ng/. Mu.L for ease of subsequent testing and calculation.

Reagent for reverse transcription of RNA into cDNA

III RT SuperMix for qPCR (+ gDNA wiper) was provided by Biotechnology GmbH of Nanjing Nodezam, the reverse transcription system is shown in Table 1-1, and the specific operation steps are as follows:

(2) Reverse transcription to synthesize cDNA

Preparing reverse transcription system according to reverse transcription instruction

1) System for removing genomic DNA

Preparing the following mixed solution in an RNase-free centrifuge tube

TABLE 1-1 degenomic System

Table 1-1 genomic system

Gently blowing and beating with a pipette, mixing, and heating at 42 deg.C for 2min

2) Preparing a reverse transcription reaction system

(1) mu.L of the diluted sample RNA was extracted, and mixed with 4. Mu.L of 4 × mix gDNA in a 1.5mL enzyme-free centrifuge tube, and then water-bathed in a 42 ℃ water bath for 2min.

(2) After the removal, 11. Mu.L of RNase-free ddH2O and 4. Mu.L of 5 × mix qRT were added to a 1.5mL enzyme-free centrifuge tube, and the mixture was water-bathed in a water bath at 37 ℃ for 15min.

(3) Then the mixture is subjected to water bath for 5s in a water bath kettle at 85 ℃.

(4) Finally, 80. Mu.L of RNase-free ddH2O was added for dilution.

TABLE 1-2 reverse transcription System

Table 1-2 reverse transcription system

As shown in figures 1A-C, the broilers of TD group and MOP group are fed with the feed which is added and fully mixed with 100mg/kg thiram on days 3-7, and a TD broiler model is established. After the TD broiler model is established, clinical symptoms of the TD group broilers are manifested by anorexia or abolition, tibial joint swelling, difficulty in standing and walking, severe people cannot walk in a standing way, and the tibial growth plate area of the broilers is obviously thickened, so that white transparent cartilage plugs which are not calcified and vascularized can be seen. The tibial growth plate index of the broiler chickens in the TD group is obviously higher than that of the broiler chickens in the CON group due to the existence of cartilage plugs (p < 0.05). After MOP treatment, the dorking in the MOP group gradually recovers the appetite, the joint swelling is relieved, and the standing and free walking ability is gradually recovered. The structure of the tibia growth plate of the broiler chicken is gradually recovered to be normal, and blood vessels in a proximal hypertrophic area of the tibia are remarkably increased. Meanwhile, the MOP remarkably improves the weight index of the tibia of the broiler chicken (p is less than 0.05), reduces the index of the tibial growth plate of the broiler chicken and gradually approaches to normal.

Histopathology further showed a significant reduction in vascular and chondrocyte numbers, a disorganized chondrocyte distribution, incomplete cell morphology with concomitant nuclear lysis and fragmentation in the tibial growth plate of broilers in the TD group, as shown in fig. 1D. After MOP treatment, the blood vessel infiltration of the tibial growth plate of the broiler chicken is remarkably increased, the chondrocytes are gradually repaired and regenerated, the arrangement is orderly, and only a small amount of mononuclear cells exist. The results show that MOP can restore adverse effects caused by broiler TD by increasing the weight of the tibia, restoring the structure of the tibia growth plate and promoting vascular infiltration of the tibia growth plate and developmental differentiation of chondrocytes.

The results of tibial ash content and inorganic mineral content are shown in fig. 2A, and show that the ash content of the tibia of the broiler chicken in the TD group is significantly lower than that of the broiler chicken in the CON group and the MOP group (p < 0.05) on the 14 th day, and the difference is not significant on the 21 st day. The calcium content of the tibiae of the broilers in the TD group is obviously lower than that of broilers in the CON and MOP groups on 14 days and 21 days (p is less than 0.05). The phosphorus content of the tibia of the broilers in each group has no significant difference. Plasma ALP, calcium and phosphorus levels of broilers in each group were further examined (FIGS. 2B-D). Plasma calcium and ALP levels in broilers in TD were significantly lower than those in CON (p < 0.05) throughout the experimental period. On days 7 and 21, the plasma phosphorus level of the broilers in TD group is significantly higher than that of the broilers in CON group (p < 0.05). After MOP treatment, the plasma calcium and ALP levels of the broilers in the MOP group are both obviously higher than those of the broilers in the TD group (p is less than 0.05), and the plasma phosphorus level of the broilers in the TD group on the 21 st day is obviously lower than that of the broilers in the TD group (p is less than 0.05). The results show that MOP can increase the ash content in the tibia, relieve the calcium-phosphorus metabolic disorder caused by TD, and increase the activity of a bone formation marker ALP, thereby promoting bone formation.

As shown in FIG. 3, the influence of MOP on BMP-2, smad4 and Runx2 gene expression in tibial growth plate of broiler chicken is shown in FIG. 3, and the expression level of BMP-2, smad4 and Runx2 genes in tibial growth plate of broiler chicken in TD group is significantly lower than that of broiler chicken in CON group (p is less than 0.05) in the whole experimental stage. After MOP treatment, the expression levels of BMP-2, smad4 and Runx2 genes of tibial growth plates of the broilers in the MOP group are all obviously higher than those of the broilers in the TD group (p is less than 0.05). The results indicate that MOP promotes bone formation through the BMP/Smads signaling pathway.

Example 2

The method for improving the antioxidant capacity of TD broiler chicken by morinda officinalis polysaccharide comprises the following steps:

step one, selection of raw materials and breeding method

Same as example 1

Step two, sampling and measuring method

10 chickens were sacrificed by cervical dislocation at 7, 14 and 21 days, plasma samples were stored at 4 deg.C, 3000rpm for 10min and-20 deg.C after sterile collection of carotid vein blood, and the activities of Superoxide Dismutase (SOD) and Glutathione peroxidase (GSH-Px) and the content of Malondialdehyde (MDA) in the plasma were measured by ELISA kit. The expression of antioxidant related genes SOD and GPX-1 in the tibial growth plate is detected by RT-qPCR, and the specific steps are the same as the example 1.

The results of the oxidation resistance related indexes in the broiler plasma are shown in fig. 4A, and the results show that the activities of GSH-Px and T-SOD in the broiler plasma of TD group are both obviously lower than those of the broiler chicken of CON group (p is less than 0.05), and the MDA level in the plasma is obviously higher than that of the broiler chicken of CON group (p is less than 0.05) in the whole test period. After MOP treatment, the activities of plasma T-SOD and GSH-Px of the MOP group are both obviously higher than that of broiler chickens in the TD group (p is less than 0.05), while the content of MDA is obviously lower than that of the broiler chickens in the TD group (p is less than 0.05), and the plasma T-SOD and the GSH-Px are restored to the level similar to that of the CON group.

The expression levels of antioxidant-related genes SOD and GPX-1 in tibial growth plates were further determined by RT-qPCR as shown in FIG. 4B. In the whole test stage, the gene expression levels of SOD and GPX-1 genes of tibial growth plates of broilers in TD group are obviously lower than those of broilers in CON group (p is less than 0.05). After MOP treatment, the expression levels of SOD and GPX-1 genes of tibial growth plates of the broilers in the MOP group are obviously higher than those of the broilers in the TD group (p is less than 0.05). The result shows that MOP can reverse the abnormal change of oxidation indexes caused by TD and promote the expression of antioxidant genes, thereby improving the antioxidant capacity of TD broiler chickens.

Example 3

The method for improving TD broiler intestinal flora by using morinda officinalis polysaccharide comprises the following steps:

step one, selection of raw materials and breeding method

Same as example 1

Step two, sampling and measuring method

On day 21, the cecal contents of 4 broilers in each group were collected, stored at-80 ℃, then iced and homogenized separately, and the microbial genomic DNA in the samples was extracted using the QIAamp DNA tool mini kit according to the manufacturer's protocol. To ensure successful isolation of DNA, the concentration and purity of the DNA extract was determined using the Nanodrop instrument to determine whether DNA was successfully isolated and the integrity of the DNA sample was assessed by electrophoresis on a 1% agarose gel. Bacterial 1696 rRNA V3-V4 hypervariable region was PCR amplified with primer 338F (5'-actcctacgggaggcagca-3') and primer 806R (5 '-GGACTACHVGGGTWTCTAAT-3'). The amplificates were purified and quantified using the aggregate AMPure Beads and PicoGreen dsDNA detection kit (Invitrogen, carlsbad, CA, USA). Constructing a DNA library, and sequencing the amplification product on an Illumina Miseq 250 platform. Alpha diversity was evaluated using the Chao1 index, the Observed _ species index, the Simpson index, and the Shannon index. Beta diversity was evaluated using Principal co-ordinates analysis (PCoA).

The effect of morinda officinalis how polysaccharide has on TD broiler intestinal flora is shown in fig. 5, and the results show that Venn graph shows the overlapping condition of OTU distribution among each group, and the results are shown in fig. 5A, 747 OUT distributions are observed in three experimental groups, 457, 537, 311 OUT distributions are respectively found in the cecal contents of broilers of CON group, TD group and MOP group, and the total number of OUT is 163 (fig. 5A). Beta diversity of gut flora was shown by PCoA analysis, and the results showed significant differences in gut flora clustering among the 3 test groups (fig. 5B). The Alpha diversity of intestinal flora is reflected by 4 indexes, i.e., simpson index, chao1 index, observe _ speces index and Shannon index, as shown in fig. 5C, and the results show that there is no significant difference in Alpha diversity of the broiler intestinal microflora of each test group. To analyze the specific changes in the cecal intestinal microflora of broilers in each test group, we analyzed the relative abundance of the dominant flora (10 bacteria before the relative abundance at phylum level and 20 bacteria before the relative abundance at genus level). At the phylum level, the relative abundance of Firmicutes in ceca of broilers in MOP group was significantly increased (p < 0.05) and the relative abundance of Proteobacteria (Protebacteria) was significantly decreased (p < 0.05) compared to broilers in CON and TD groups. The abundance of Actinobacillus in cecum of broiler chickens in TD group is obviously higher than that of broiler chickens in CON and MOP group (p is less than 0.05). The relative abundance of bacteroidetes (bacteriodes) was not significantly different between groups (fig. 5D). On the genus level, compared with the CON group broiler chickens, the TD group broiler caecum has a significantly increased abundance of beneficial bacteria such as Lactobacillus (Lactobacillus), blautia (Blautia), ruminococcus (Ruminococcus) and segmented filamentous bacteria (Candidatus _ Arthromitus) and intestinal membrane barrier, reduces inflammation, provides energy and immune function related to the organism (p < 0.05), aerococcus (Aerococcus) and Corynebacterium (Corynebacterium) cause bone and joint infection, and causes osteomyelitis and suppurative arthritis (p < 0.05), and the MOP group broiler chickens (Blautia), rumen (Ruminococcus) and segmented filamentous bacteria (Candidatus _ Arthromitus) have a significantly increased abundance (p < 0.05), and the group broiler chickens (Blautia), rumen (Ruminococcus) and segmented filamentous bacteria (Candidatus) have a significantly increased abundance of beneficial bacteria such as Lactobacillus < 0.05), aerococcus (aoccus) and Corynebacterium (Corynebacterium) have a significantly decreased abundance (Corynebacterium) and Corynebacterium) as < 0.05. The result shows that MOP can improve the intestinal flora structure of TD broiler chickens, improve the abundance of beneficial bacteria, reduce the abundance of harmful bacteria and further regulate and control the metabolic function of organisms.

It should be noted that the above-mentioned embodiments are exemplary, and that those skilled in the art, having benefit of the present disclosure, may devise various arrangements that are within the scope of the present disclosure and that fall within the scope of the invention. It should be understood by those skilled in the art that the present specification and figures are illustrative only and are not limiting upon the claims. The scope of the invention is defined by the claims and their equivalents.

Claims

1. A method for relieving tibial cartilage dysplasia of broiler chicken is characterized in that morinda officinalis polysaccharide is added into broiler chicken food.

2. The method of alleviating tibial dysplasia in broiler chicken of claim 1, wherein thiram-induced improvement in tibial cartilage development in broiler chickens is induced by morinda citrifolia polysaccharide through BMP/Smads signaling pathway.

3. The method for alleviating tibial dysplasia of broiler chicken of claim 1, which is characterized by the following steps:

step one, selection of raw materials and breeding method

All broilers were housed in standard room temperature and relative humidity light/dark cycle facilities and provided free of water and food. The experiment randomly divided the broilers into 3 groups of 10 chickens each, each group was 4 replicates, respectively control group (CON), thiram induced TD group and MOP treatment group; on the 4 th to 7 th days, the CON group is fed with normal diet, the TD group and the MOP group are fed with diet added with thiram of 100mg/kg, on the 7 th to 21 th days, the MOP treatment group broilers are added with MOP of 500mg/kg in drinking water, and the test is carried out for 21 days.

Step two, sampling and measuring method

The broiler weight was recorded daily during the test period, 10 chickens per group were sacrificed on days 7, 14 and 21, respectively, by cervical dislocation, and plasma samples were stored at 4 ℃,3000rpm for 10min and-20 ℃ after carotid vein aseptic blood sampling. Measuring the length of the tibia and the width of the growth plate and performing H & E staining; taking the right shin bones of 6 broilers in each group, and measuring the contents of ash, calcium and phosphorus in the shin bones; plasma calcium, phosphorus and ALP levels were determined by the kit.

4. An application of radix Morindae officinalis polysaccharide in relieving tibial cartilage dysplasia of broiler chicken is provided.

5. A method for improving the oxidation resistance of broiler chicken with tibial cartilage dysplasia is characterized in that morinda officinalis polysaccharide is added into broiler chicken food.

6. An application of radix Morindae officinalis polysaccharide in improving broiler oxidation resistance of broiler with tibial cartilage dysplasia is provided.

7. A method for improving intestinal flora of broiler chicken with tibial cartilage dysplasia is characterized in that morinda officinalis polysaccharide is added into broiler chicken food.

8. An application of radix Morindae officinalis polysaccharide in improving intestinal flora of broiler with tibial cartilage dysplasia is provided.